Iowa State University Capstones, Theses and Retrospective Theses and Dissertations Dissertations
1945 A cytological study of the costal marrow of the adult horse and cow M. Lois Calhoun Iowa State College
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Recommended Citation Calhoun, M. Lois, "A cytological study of the costal marrow of the adult horse and cow " (1945). Retrospective Theses and Dissertations. 13465. https://lib.dr.iastate.edu/rtd/13465
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fhssis Eubscitted t0 tbft Sr& E0CT08 OF ?H1I.Q£0?H¥ Msjor Subjsets lf®t«risRrx iaatoKy Signature was redacted for privacy. la Chsrge of Mejor Work Signature was redacted for privacy. fejor Bepftrtasat Signature was redacted for privacy. Dssis 0t OTf'duEt® Goliegt lose St»te Colissg© 194S UMI Number: DP13152 UMI UMI Microform DP13152 Copyright 2005 by ProQuest Information and Learning Company. All rights resen/ed. This microform edition is protected against unauthorized copying under Title 17, United States Code. ProQuest Information and Learning Company 300 North Zeeb Road P.O. Box 1346 Ann Arbor, Ml 48106-1346 / /' • • Table of Contents IKTRODUCnOH 1 HESTOHY 1 LITERA'IORE REVIE® 3 GROSS MD MICROSCOPIC MATOMY 01' BONE MiMfflW 7 MTlRIii. MD METHODS 9 Obtaining the Samples 9 Oxalate Tubes 13 Staining end Counting Technic 14 RESULTS 18 Blood Studies 18 Bone Marrow Studies 24 Terminology ...... 24 Myelograms S7 Description of cells , . S7 Mitosis 39 Myeloid-eryttroid ratio 40 Bone-healing process 41 Sempliag experiment . «... 41 DISCUSSIOM 44 . 63 LITEHATOEE CITED 66 iraOlflLEDGMMTS 86 PBOTOGBiSHS MD LMDS 87 r7^76 1 liJTBOIDCTION Importeat contributions hP.Te been mede to the microscopic anatomy of bone marrow in men, monkey, and ©aall laboratory snimels. A seercli of the literature failed to reveel eny similar reedily available studies on fe.mi gnimals. In view of the increasing importance of bone merrov; as a dlegnostic agent, and as s contribution to a field of histology relatively unexplored, this atudy of the bono marrot? of the horse and cow was under taken. fflSTORS: According to Michels (19318), Jen Swainraerdjaii first observed the red blood cells in 1658. Wiseman (1934) stated that hematology was first established as a branch of scientific medicine in 1770, but it was not until 1838 that Weber first saw nucleated red blood cells in man. Scott (1959) found that Eobin described nucleated cells of the red marrow in 1849, His torians such as Sabin il9S8), Michels (l931a) and Scott (1939) agreed with Strieker (1870) that Neuman (1868) was the first to associate blood forma tion vjith bone marrow, that Bizzozero (1868) confirmed his prork that seme year, and thet Claude Bernard reached the same conclusions the following year. They ell vioriced on human end rabbit msterial. According to Wilson (194:2) the bone marrow picture in pernicious sn®iila was first described by Pepper in 1875 and Cohnheim in 1876, Dosn (1939) felt-that real progress in hematology began in 1891 when Ihrlich reported the effects of specific dyes on the blood cells, Sabin (192£) imputed the fonaation of our knowl 2 edge of blood to Ehrlich. Wolff (1903) performed bone msrrow punctures on experimental animals. According to Scott (19S9) and fiintrobe (1942), Pianese executed a human marrow biopsy ia 1903. Shedini (1910) conducted similar clinical biopsies on bumsn subjects in 1908, These workers end their followers all used tibiel or femoral marrow end it iras not until 1923 that Seyferth (1923) first introduced a sternal trephine method. Arinken (1929) followed with the sternal puncture technic which hss become so popu lar 8t present. Classic among the early WDrks on bone marrow are: Pappenheim*s (1899) article which deelt vdth a general comparison of one type of bone marrow cell isith enother in various ages, using rabbits and one dog; Domi ni ci's (1901) papers on the straoture of the hmetopoietic system of msri- mels; DaatschsJsoff's (1908) thesis on the development of blood in the bird; Maximow's (1910) investigations including the embryonal histogenesis of the bone marrow of memmals; i'errete's (1916) treatise on hmatopoiesis end Sa- bin's (19S2) dissertation on the origin of the cells of the blood, 35'urther Eiommental publications on the subject of bone marrow include those of Schilling (19S5), Iskanazy (19S7) and Sabin (1928). Gilmour (1941) made a r^narkable contribution to the general subject of hujiian intra-uterine and neonatal heimatopoiesis. Plum (1936) said that Rudolph Wagner attempted to count the fomed elements of the blood in 1849 but that the first real count was made by Vierordt in 1852, Garrey and Bryan (1935) claimed that Nasse had been in terested in blood counting prior to 1842 and had •written a section on the subject in Wagner's "Eandworterbuch fier Physiologie," Plum (1936) attribu ted; the counting chamber to a Dutch investigator named Cramer working in 1855, •vsltli further modifications by Gower in 1877 end Mferow in 1884. In 1854 Welcher was using distilled water es s diluent for leucocyte counting •fa-ut Ttoma later discoTered that the addition of acetic acid reduced the emount of water necessary (Plum, 1936)< Potain introduced the pipette in 1887 (Garrey and Biyen, 1935), These seme historians credited the dexrelop- ment end precision of the pipette and cross-ruled chambers to Welcher in 1854, Abbe in 1878 end Lyon and Thoma in 1881. LiTERAi'usi: mnsn The study of hematology has only recently come to the front in veterinary medicine, Wittmann (1988) stressed its importance in the field of animal disease, veterinary surgery, and animal breeding. Hovfever, as early as 19Q6, Zunta, et made histological studies on the bone marrow of dogs at sea level, and on animals acclimated to higher levels, Notable contributions on the subject of nonnal monkey bone marrow have been made by Suarez, Diaz-Eivera and Eernandez4lorales (1943) and Stasney and Higgins (1936). Science is indebted to Alexendrov (1930), Fairmsn and WMpple (1933), Stasney end Higgins (1937), Mulligan (1941, 1945), Van Loon and Clark (1945), Iberl (1943) and Bloom and Meyer (1944) for studies on the bone marrow in the dog. In the first decade of this century .Kulbs (1909) worked on the development of bone marrow using the dog es en experisaentsl animal. Bloom (1944) compared blood and bone marrow in relation to pyometre in the dog, A variety of studies has been carried out on rabbit bone marro'# including the work of Lim, Sarkar end Broivn (1982) which entailed the estab lishing of the normal histological picture in a study of the effect of feed 4 ing thyroid, Sabin end Doan's (1927) worlc oa the bone marrow end blood in developing rabbits by Ssbin,^ (1936), the lymphocyte content by Yoffey and Pernell (1944), the distribution of bone marrow, bone and bone esh (Diets; 1944), end Jordan's (1920) study of the giant cells of the rabbit and guinea pig bone marrow. Sundberg end Bomiey (1942) compared the lymphoid cells of Ipiph nodes and bone marTOv? of rabbits and guinea pigs. Epstein end Tompicins (1943) used the guinea pig es an experimental snimsl in e com parison of techniques for the different!el counting of bone msirrow cells, louriquand, Hevol and Edel (1944) chose the feaiur as the site for e study of the myelogrem in nomal guinea pigs. Similar studies were mede by lair- man end Corner (1934), Milmsn, Listengarten and Surbanalievf (1934), Stesney and Higgins (1935), Kindred (1940) and (1942), Plum (194S) end Indicott and Ott (1945) on the albino rat. A noraial bone raerrow study on white mice xvss made by Petri (1934). Hammon and Enders (1939a and b), La-wenoe ^ a], (1940) and Eiser (1943) studied the bone marrovi in connection with infectious pan- leucopenia of cats, Corradetti (1934) studied the bone marrow of new bom cats in connection witii some work on the blood find bone merxow of healthy human beings, Gutig (1908) made e survey of the bone marrow in sisine while studying the morphology of the blood of the species. Do?a).ey (1915) used the adult guinea pig to determine the origin and development of both the eosino™ phil and the "hesnatogenous mast cell," Barbieri (1935) studied the altera tion of the bone marrow ia association with hepatic distomoniasie in sheep, Kingoen (1921) investigated the bone marrow of the sheep and pig to trace the origin of the eosinophil. Other investigations which serve to stress the grovdng importance of the bone marrow as an experimental organ and the need for knowledge of the noimal, include such irorks as the following; Beilicke (1958) observed 5 the effect of iron on the blood end bone marrow of rabbits; Cestrodele, jJ, (1941) used dogs to study the comparatiTe effects of estradiol and stil- bestrol upon the blood, liver end bone marrow; Menkin (1945) studied the effect of the leucocytosis-promoting factor on the femoral bone marroisr of dogs; Isntrie (1934) investigsted the cytology of the erythroblast in the gro\?ing guinea pig embryo; Saith and Hastings (1935) made a study of the megekaryocyte and blood platelet of the rat; Dougherty, iilliems, and Gard ner (1943) reported the changes in the myeloid and lymphoid tissues of estro gen treated dogs; temperature variations between centrsl and outlying bone marrow of the rabbit, pigeon and elbino ret were discussed by Huggins, Block- sam and Hoonan (1956); Huddleson end Hunger (19S7) gave tbeir attention to the phagocytic activity of bone marrow cells isith the guinea pig as an ex perimental anima^.; potter and Ward (1940) investigated the development of the megekeryocyte in adult mice, end Nettleship (1942) discussed the rabbit-bons- marrow changes produced by specific antibodies, Likeiijise Holderlin (1938) dealt ^vith the bone jasrrow and blood picture in the sensitized rebbit. The effects of sulfsnilsmide on the bone marro'S^ of rsts wes presented by Higgins and Mechella (1939), Damede and Leger (1939) gave the cellular percentages in bone marrow following experimental anemia in the rabbit. Little iffork has been done on the horse and cow. Due to vrer condi tions and failure to receive foreign journals part of the work that has been done is not available, Ickemecht (1912) was probably the first to work on the horse. He spoke of the gross changes occurring •(d.th age and gave a brief description of the marrow cells. Yericak (1935) studied the marrow of n bones of trunk of horses, cattle, swine, dogs and cats, Hjarre and Berthel- een (1938) presented detailed cellular counts of the bone marrow of ten nos> 6 liorses ead compared that with, a similer count in horses v {l941a) on the noraiel bone marrow. The sems year Msrcato {1941b) made a study of bone marrow in boTine fascioliasis, Mitchell (1940) discussed hy perplasia of the bone marrow and osteohemetochromatosis in a yearling steer and steted that the bone marrow "exhibited histological changes," lo noimel picture was given, Stssney end Feldraan (1938) mede hemetologic and histo logic studies of the bone marrow from the femur of a calf with leuksmic lymphoblastoma but again no nomal was used for comparison, Richter (1938) in discussing leucemia in animals said of cattle-that lesions were not found in .the bone marrow. According to Jaimai (1954) histologicel investigations of the bone marrow of the cow in leucosis were made by Indres {19S1), Leng- wenant (1931) end Tollner (1931), Hbvfever, they did not mention eny his tological studies being made, Jamai further stated that in the literature reviev; of leucosis in the horse, "bone raarro?; was seldom mentioned perhaps because no variation was apparent," Ellenberger (1921) gave a brief resume of the bons mairow in domestic animals with scent reference to any specific animal. *Hot available 7 GROSS MC MICROSCOPIC MATOiilY Of EONE immW The medullary cavity of the bones of the young of all species is filled isith red marrow^ Ae age increases red bone marrow is gradually re- plctced by a labile,yellovfc',fatty marrow until in the adult, red marrow is coa- fined largely to the axial skeleton. According to Ackerneckt (1912) red mar row of the adult horse is confined to the proximal ends of the femur and hu.-- rerus, the pelTic girdle, Tertebrae, ribs and sternum. Yariwk (1935) studied the bones of 100 horses and 300 head of cattle inacroseopicelly. He found thet the deposition cf fatty raarrow occurred much earlier in cettle than in horses. This pi-ocess started in the axial skeleton as esrly as 9 months in cattle. While he detemined no definite ege for the transition from red to yellow marrow in horses he rerely obserred fatty aress in the axial skeleton of any horse under 8 years of age. In e study of the ribs of a 10 yesr old horse,¥aricek observed that active msrrow was present for e distance of 20 cm. in the vertebral end, that the central pert had both red end yellow laer- row, while the sternal end was filled with fat. The 14th rib of the same animal had only a little fatty marroiv in the ventral end. In one 50 year old horse no fatty marrow wss apparent macroscopically in the ribs. Tari(^ek TOuld explain the few granulocytes end the large numbers of lymphocytes in the blood of cattle by the large amount of fatty marrow. Some of the thoracic vertebrae were completely filled vdth fatty marrow. As in the horse, the dor---- SBl eindsof the ribs retained active marrow. The central part had both red end yellow marrow and the sternal end became a reservoir for fat. The sternum too had areas of fatty marj^ow in it. By microscopic examination he observed that some of the fatty raarrow centers contained foci of red marrow. Trautmaa and 8 Fiefciger^liggl) found gelatinous marrow in older horses, Taricsk (19S5) did not find geletinous marrovf in hesltby, active,aged horses. According to CJowdry (1942) gelatinous marrow should he expected in very old people. By studying the tibia, feniur, rib, sternum and vertebrae of man, Custer and Ahl- feldt (1932) ascertained that the cellulsrity of red marrow deereeaed vdth Kd- Tsncing years. Specific gravity of bone merraw ms found to be only slightly more than, 1,0 (Yoffey snd Pemell, 1944), Sisson end Grosaaen (1938) attributed the blood supply of the long bones to the large medullary or nutrient artery which enters the nutrient foreaion, extends through the canal in the compact bone and ramifies in the marrow, A satellite vein follows the opposite course, Eoan (192S) found that periosteal vessels along the shaft sM some of the vessels nes.r the ex tremities furnished additioncd blood supply to the bone marro'ss'i The venous drainage corccesponds to the arterial supply. According to Eban (19SS) the thin welled venous sinusoids making up the vasculer bed of the marrow ere the most characteristic feature of the gross circulation. He and Eoan, Ounning- hem and Sabin (19E5) contended thet the vascular systm is s closed system. Osgood and Secjnen (1944) described the human msrro's'? as the "Isrgest, most 'isidoly dispersedj end least homogeneous orgeja in the body" with a volume one or tm times that of the liver, Cb-wdry (1944) stated that msrrow malces up about 5 per cent of the tot&l body weight, Mechanik (1986) gave the weight of the bone marrow as 3.4 - 5.^ of the body weight and estimated 5/9 ga, of marroTT to 1 gm, of blood, Dietz (1844) gave some interesting statistics on the relative weight of rabbit bone marrow; 1/3 the "weight of the skeleton, 8,^ of the total body \?eight, 2/5 the -sleight of the liver end 50 times thet of spleen. *See Sllenberger (1931) 9 According to Piney {192S), Trautmsn and Fiebiger*(l931) and Wolff (1933) there are no lymplmtics in the marrow, Fischer (1917) found lymph nodules in 58 out of 61 humen cases investigated. Lymph nodiiles were con sidered by Mayer and luruta {1924} snd 'Williejns (1939) to be e normal but variable constituent of humt-m marrow. Trautmsn end Iiebiger*(l9Sl) credited man snd eat marrow xvith lymph nodules, Wolff (1933) did not find nodules with any regularity. Drinker and Yoffey (1941) concluded thst "follicular accumulations in the mtrroiv do not seem to be true lymphoid nodules isith cells showing active division," Apparently the nerve supply in men is confined to the walls of the blood vessels and no ganglion cells have been found (Wolff 1933). Acker- neckt (191S) found nerve bundles in the humerus of the horse. MATMliiL MD MSTIDCS Obtaining the Semples The cattle used in this investigation consisted of 13 cows snd 1 bull in a herd used by the Department of Veterinary Obstetrics at Iowa State College. The horses were a miscellaneous group of animals, five of which TCre brought in to the Department of Veterinary Anatomy at lora State College and the other tiso to the clinic at Miohigsn State College, All the horses were old but apparently free from disease as far as could be determined by general appearance and blood determinations. The first problem Vfas to determine a satisfactory place to secure a marroTif sample. Since both the horse £ind cow have a heavy musculature cover ing the steruum, it seemed wise to trj"- to find an ares more accessible, Hjarre and Berthelsen (1938) contended thst a sternal puncture was easily exe cuted in the horae and they did not observe any complications from the proce sses Ellenberger (1951) 10 dure* The animel ms confined in a prone position and an especielly constructed trocar wes driven into the middle or back steraebra by s "light" tapping with a hard rubber hismer. Ia any sternum, that was ezesrdned by the euthor no light tapping would succeed in penetrating the wall of the stemebrae. jileo to here to confine such large enimsls in a lying position seeined a distinct disadTCnt- age. According to Tsridek (1938) the stemxim of the horse was not suitable be-" cause of large muscles end such a procedure in the cow was 8wkt?ard end time consuming. According to Ickerneekt (1918) the red marrow of old horses is con fined to the stemuia, ribs, fortebrae, proximsl ends of the femur and tibia jsnd to the ilium. After several unsuccessful attempts to obtain marrow from the fffiiiur, ilium end mendible of the horse, the ribs were chosen as the best site. Plats I, i'ig. I shows the generel area end Pl&te I, Jig. 2, illustrates the fact that parts of the 8th to 18th ribs are relatively exposed in thet region being covered only by skin and fascia. It is well to go as high es possible and still avoid the letissimus dorsi (k) and serratus posticus (m) muscles because it seamed to be difficult to obtain sufficient merrow more ventrslly. The seme technic was applied to the cow using the 11th, ISth or 13th rib, (Plate II, ilgs, 1 and E) She animal may be confined in a stoclc or restrained against one side of a stell. Little or no resistance to the operetion wss or dinarily encountered. It is advisable to brush the beck and side of the aniiaal with B grooming brush. Wiping the surrounding eree, particularly ebove the prospective operative site i^lth a dsmp cloth msy also aid ia removing some dust and particles which might leter fall into the open wound. The general eres was pelpated until the rib •which hed the least emoxmt of fsscia covering it •was located. The chosen site was shsved or the hair clipped closely, the area was TO^ed with a soap solution and iodine applied, A local anesthetic such as 2$ 11 procaine hydrochloride ;vas next sdministered. The skin wss enesthetizsd first, the underlying fascia next and finally the periosteum, using about 10 cc. of the procaine solution. After s few minutes a short incision was made in the skin and then the fascia end periosteum "vvere incised, A No. 487 Goodell- Pratt* hand drill as shorn in Plate III, equipped with a straight shank S/SS" johlters* drill, "stfss used to bore into the marrow csirity. 1 point midway be tween the anterior and posterior borders of the rib should be chosen for in sertion of the drill because there is danger of missing the merrow cairity com pletely if the drill,goes through either border. Such an accident would entail the danger of penetrating the thoracic cavity. The drill "gives" when it hits the marrow, so it vraa not difficult to sense when the marrow V&B reschedi The drill wfis removed from the rib and a cannula with stilet -Rlth the seme outside diameter ss the drill was inserted into the drill hole. 1 Jen-Sal needle tro- cer, J. S. 39121 in the 1940 catalogue** wfis used. The stilet was reinoved amd an air tight 10 cc, syringe attached to the cemmla. One ec, or less of mar row was then drawn into the syringe. The syringe wes separated from the cennule and the marrow ejected in to en oxalate tube to prevent coegulatioa (See page is). The tube Vv-es held in a horizontal position and tapped to mix the msrrow and oxalate. After obtain ing the sample the cannula was withdram, iodine applied end the incision left open. Healing took place rapidly and after the heir had grovsn out the site of the incision could not be deteHrdnsd. Ysrio'ek (1955) found that trephining the rib need not leave a scar. With the exception of the necessity for drill ing through bone and the use of a cannula and stilet the above procedure ma *6oodell-Pratt Company, Greenfield, Mass. **J"en-Sal Laboratories, Kansas City, Missouri, 12 patterned after that of Osgood and Brovmlee (193'!') for sternal puncture in man. All the necessary instruments end supplies 8re illustrated in Piste III. There has been some criticism of 'withdrawing more then 1 ce, (Isaacs, ^ 1940} on the basis that the ectual marrow %lthdrami is too diluted with circulating blood. Jaff^ (1936) suggested 0.1 or O.g cc. Ac cording to Hjarre (1943) the sample should be more Tiseous theai blood end greyish red in color. It is possible to eater s large sinusoid and withdraw material that differs little from peripheral blood, (Jones 1940 and Hjarre 1943). Should this occur, the procedure should be repeated at e different level or on a dif ferent rib. Slides were taksn to the operfitive site in event fresh smearB ?f0re ever desired. The state of the marrow can only be deteiroined after having enalyzed the blood picture (Sabin 1923). Msnaugh (194£)) suggested that correlation of the marrow picture ifdth that of the peripheral blood might eventually result in the ability to read the laarro?? hanogram by the blood findings alone. Blood ssmples were obtained prior to procuring the marrow es the lat ter procedure might excite the animal enough to alter the blood picture. The jugular vein was chosen as the logical place to obtain such a ssiaple. I'he blood was directed into en oxslate tube and sheken well. The usual aseptic venipuncture technic for lerge enirasls was followeq. Conner (194S) described this technic. The samples were taken to the laboratory end blood and bone marrow smears made immediately. Then total red and white counts v.'ere done on the blood and the amount of hemoglobin determined, Osgood's (19-^) technic was 13 followed. Totsl red ond nucleated cell counts on the msrrow semples were purposely oroitted because there was such e large vsriatlou due to dilution Ti5ith circulstiag blood that the dste seemed to have little or no value. Nordeasen (1935) and KoUdel and Loroy (1939), workisg with human bona marrow, found this to be true slso. The above technic was defdgned for the living aniuiel end givee the optijauffi noiKsd cytological picture, Eo'^^ever, the occasion may present itself that postmortem-marrow exesnination TOuld be necessary, lintrobe (194E) felt that the ssanple should be secured vsithia 2 hours after death to get good prep arations as autolysis occurs soon, Rohr and Hsfter (1937) made a special study of postniortera changes in human, bone marrow. They concluded that death had little effect on the myeloid elements, but the erythroblast matured •<h a subsequent change from polychromstie to oxyphilic forais end extrusion of the nuclei. They also observed a, decresse in the number of neutrophils beginning a few minutes postmortem. The nuclei of the remaining neutrophils swelled and beceme vB.cuol8tea after 2 hours, Ozalate Tubes The blood TOS collected in ordinary test tubes. Dunham fennentatioa tubes were used for the m&rrow because they were short snd had a relatively aiall bore. Consequently ths marrow and oxalate could be mixed more thoroughly. The oxalate tubes were prepared by evaporating to dryness 0.1 ce. of a 2$) potassium oxelste solution for each cubic centimeter of blood or msrrow (2 Kg. of dry potsissium oxalate per cc.). The requisite amount of oxsilate solu tion was measured, into the tubes end they were pieced in e. drying oven. Maurer and Jones (1943) evsporated the oxalate solution st e tsapere- ture belo?/ 80° C. since higher temperatures converted some of the potassium 14 oxalate to carbonate end consequently coegulation v/as not inMbited. Too much oxalate tended to distort the noraal cell structure. The cells sppeered shrunken and the staining was too intense. Osgood, Easkins and Trotman (1931) rscosiaended oialated blood be- cauce of greater conTenience and belisved its use resulted in e. higher de gree of accuracy since the sBmple was larger, moi-6 time could be tsken end duplicate estiiaations could be ran. They suggested £! 24 hour tiii).e liiait for heraoglobin readings and total red and white blood cell counts, and a one hour limit for the sniear differentiel. In some inTestigations at the University of Mnnesote., Kernkesap (1948) concluded that if osal&ted blood samples were examined vdtMn 60 to 90 minutes after obteining the blood, few changes re sulted. Me.urer and Jones (1943) agreed v?it-h tMs view. Staining end Counting Technic The slides vfere soaked in sulphuric scid-potaasiuffl dichroraate clean ing solution 24 hours, rinsed in running tep water for ¬]ier 24 hours, rinsed in distilled water and stored in alcohol. They were dried as needed, A 24-gauge bacteriology platinum KLRE loop WES used to remoye the blood end marrow from the tubes in making the smears. To prepare the smears, the drop of blood or marrow was placed on the right end of a clean islide. A' second glass fslide, resting at an angle of about 45° \?ith the first one, was moTed along the first slide from the left until contact of the second slide TOs made ulth the drop. This junction spread the blood out slong the -width of the slide. With the second slide still st the sane angle, it was pushed to the left and the blood spread out in a smear behind it. The smear •»as waTed in the air to insure rapid drying. In a properly prepared smear the cells should not touch-and the end of the smear should reflect rainbow colors. 16 to stain, tfe® saieer wes mtirked off Kitli s tbx oeEcll tsiais requir ing 6 miiiistj® of at?dnlng uolutlon. Wright^s stsiniog tecliaic ss outlined by Osgood SBd Ishworth (1937) ms foliowsfi. OHS-tsEth gm. of Wright's dry gtsia wfiS jaixed with BO co. of acetons-frse tsethyl elcosol siid left over sight before U!?e, A buffer solution %lth & pS of 6.4 wes prepuraj? by Sia- solvisg 6.5S gtjis. of ssoaopotassim pliosphst© sad £.56 p\6. of snliyarous di- sodiUB. phosplmts in e Iltar of CiaUllsd v/cter. One QC. of chlorofom ma fidded. Six to 12 drops of thsi 6ye, depesdicg on. the aize of the sseBr, tfers left oa the slice for two miauteg, then diluted edtb sn 8c,ua:i number of drops of buffer and left seven Mautes longer. With tls® Kiide ia s iiorizoEtal posi tion tii© staining solution ??ss rijised off sad ttse slide WiBhiSfi for tMrty ssconfis tsit,li a brisSs: straem of running tsp wtor. fhe. slides were eir dvim, Ho cover slip •^.-ss applied. Xastead the smear eoTOrec utith a tbia fiisn of isiKiersion oil fit ths tise of es/fjisinctioa. In this stix&j the fillsd aosjating cfesKbero were sxcsiaod with t53.8 low po'ger objecti-ve for gsnercd uaiforaity of cilstriMtlon, bubbles, or aay foreiga SBtsriel. If th& iaitisl survey no iaequfiiity ot cell \iis- persioQ, babfeles or other artefeets the counts tfsre rasds. Diffsreaeaa sx- ceediag the "atfengKrd liislte'* wer© not ciscredited. Both sices of the cte- bsr wore counted er,« totals wrs gsteisninsfi frojs tjsosa figures. Hemoglofeia resdiEga wers aecie on a Dcre h^aoglofeinoiceter. Altfeougb this instrussnt woald not be precis® eaougb for resesrob speoificidly oa hmo- globiE it sss ^leeasd atfficieifitly sccursts for use iere. Pium (1936} if£S of the opisioa thet the wefual fiiatributioa ot the cells in thg blooti sae&r pleyed tho pre^ominaat role is tfeo error of iadi- vl rfe.y (194-S) recoKKonded counting 200 ox 400 white blood cells to re duce the error of rr-ndoa ssmpling. He found fui'ther thet the choice of area in Vfhich. to count the cells did not play as important part ss thought by ear lier investigators. With the low povfer magnification s field was chosen in T(hich there vjas fin even distribution of nucleJited cells, the red cells did not OYerlap end staining was:sharp. The edge of the smear was evoicled. In the blood mears 200 cells were coimted. at rsndom by going back pM forth acrose the smear. A Spencer research mici-oscope with widefield 9x oculars, and 16, 8, 4 end 1.8 mm. otjectiTSS iBas.used. All differfjntial counts were made vdth the 1.8 am. oil immersion objective, magnificstion 95x, meJcing a magnification of 855x. A Spencer lamp model 370 tos the source of light. A 100 watt bulb was customarily used but for cytological detsils a 200 or even e 300 wstt bulb vdll bring out spacisl structures. Thorns pipettes were used throughout for diluting the blood for total counts. The Heubauer "Bright Line" counting chsmber wes aaployed. Separata counting chembers were used for the red and white counts. This avoided any possibility of carrsring over tny of the acetic acid used in the white count to that of the red count. Standard textbooks on hemstologicsl technics cej.l for e variation of leGs thsn 18 to SO cells between the 5 groups of squares for the red count and 17 Trsriatioa not to ezceed 8 or 10 cells 'bet^^eea scueres in the- ^Mte count. This Tiould be v.ltM2i limits of 180,000 to 200,000 and 160 to 200 cells re- spectiToly for totsi counts. VJindle, Sweet sad White-head (1940) accepted ta agrsement of tv,'o eoimts•7,1 thin limits of 100,000 for the rod hlood cells and "50 for wMte blood cells. Berkson, Mageth, aad Hum (1955) trnd Hageth, Berk- eon end Hui^ (19S6) shov/ed that such stringent llffllts Ere not stEtisticelly prolsable. To the controry they found that greater differences then those v.'ould be normally ezTiected in frora 50 to luoro than 90 percent of the eoiints, OMson (1945) KiBcle a study of sampling methods la connection ?.lth erythrocyte deterrainations. From data obtained ?Jith hGemccytosieters th&t presented e uai- foral distribution of cells &nd no bubbles, the follovdng cQnclusions were dra-vin: (l) eignificant differencee ocevirred betv;een sub,1ects; (E) differences border ing on significance Mght occur between pipettes; (3) T.lth the trained tech- nicieai there was no statisticElly significant difference betv?een successive counts on the srae ssmplis oYon though diffprenoep sj^ceeding £0 cells existed bet-weeai the 5 groups of squares. In the differential cell count of the marrow 500 cells were enumer ated. Stasnsy snd Higgius (1925) (1939), Osgood (1937), Eingery, Osgood and Illge (1937), Pitts and Padchem (1939) and Bloom and Meyer (19M) enumerated 500 cells. Wintrobe (1942) stated that 500-1000 should be counted. Davidson, D-stLs snd Innes (194E) suggested observing 400 or 500 celle, Hjarre (1943) counted 800 cells end Wilson (194S) and Thomson (1944) counted 1000 cells, Reich, Swlrsky and Saith (1944) did not do differential counts "oMng to the inaccuracy of the method." In a subsequent personel ooromunication Dr. Keich said that the inaccuracies that they found might not exist in this vitsrlc. 18 According to Cowdry (1944) a bone marrow ameer will contain endo thelial cells from the eapillaries, some reticular cells, and connective tissue fibers in addition to the developing blood cells. Cells are frequent ly injured and the cytoplaaa may have been torn and the nucleus drawn out in strands. Other cells appear to be dying. These injured and dying cells have been given various names such as degenerate, smudge, smear, or basket cells. Since ^pies (1939) considered theae cells to be "largely fortuitous and mere ly distort the picture if they are included in the differential count" and Bloom (1943) thought them artefacts and not necessary to include, it was ds- cided to omit them from the differential count. EISDLTS Blood Studies Miller (1938) made 81 determinations of the blood volunie of cattle. The average quantity of blood per pound of body weight was 27,07 cc, in in crease in blood volume occurred in pregnancy, Wirth and Mader (1988) and Zemlji^ (1955) stated that the ox has a lymphocytic blood picture,* They characterized oz blood as having azure gran ules in the lymphocytes, ocoasional stab cells, small eosinophilic granules, and commented on the especial size of the neutrophil. According to Gkjodall (1910) the general morphology of oi blood does not differ greatly from human blood. Du Toit (1916) also found ox blood similar to human blood except that it had a high lymphocyte count and a low neutrophil count when compared -Jfith man, Gudim-Lewkowitsch (1929) in a study of the differential number of the neutrophils in 15 normal cows gave the follo^dng data: juveniles, 0,S^; stab, 45,'?^52-lobed, 35,^;3-lobed, 14,9^; 4-lobed, 2,9^; end 5-lobed, 0,6^, Zeml- jiS^'s (1935) results on 50 animals differed somewhat with 12.3^ juveniles. •Majority of white blood cells were lymphocytes. 19 21.6^ stab cells and 65.1^ segmented, Eraser (1930) found 0.1^ juveniles, S.SJJ stab and 96.3^ segjaented. Besel and Levek (1928) stated that there were no Bjyelocytes, 0.1^ juyeniles, 2.8^ stab cells and 41.^ segaented neutro phils In nomal ox blood. Kennedy eind Climsnko (1931) reported the normal imeth count for tlxe cois as 2.34 (weighted mean). Goodall (1910) characterized the neutrophil of horse blood as haying aa unusual degree, of lobation and especially fine granules. He found the ^siaophil to contain large spherical or OTOid granules. V/irth (1931) men tioned the large eosinophil, large basophil, lai^e platelets end.preponder ance of neutrophils in the horse. It seems feasible, since any so-called "normal" limits of either blood or bone marro-ar must be arbitrarily chosen, that some animsls without evidence of abnomality may hare counts that do not fall within the range ordinarily considered nomal. Shukers, Langston and Day (1938) were of this opinion. Tables 1 and 2 sufflmarize the literature on blood studies of cattle and horses. Tables S snd 4 present the hemogrsms of the experimental animals used in this study. Table 1 Heraogrems reported on cettle Hsmogiobin Rontent*** R.B.C. IV.B.C. Differentia 1 count, ner cent Mo. of Gm.per millions hundreds Weutro- Lympho Mono— JSosino-- Beso- Autlior ^nimsls ^ 100 cc. "oer cu.mm. T)er cu.mm. Dhils cytes cvtes ohils phils Bimo ck & Thomp son (1906) ISF 59.75 8.25 6.1526 54.86 30.49 54.22 1.47 13.15 0.59 DuToit (1916) 7F 6£ 8.5 6.54 78.6 38.8 49 3.7 8 0.5 Ktihl (1919) 24 65 10 5 0-1 Meyer (1924) 44,3 46. 7 5.3 3.7 Basel & l.ewek (1988) 44.6 44 1.6 9.6 0.2 Kohanawa (1928) 12 6. 779 82.1 33.0 51.7 4.3 10.9 0.1 Sergent ^ (1929) 39 10.0 28 59.5 7.5 10 1 .0 Canham (1930) 1£F dry 6.66 97 34.0 53 4.0 8.0 1.0 Fraser (1930) 41 28.4 54.7 6.7 9.9 0.1 Scarborough*(19S1-3S) 60 8.£6 6.6 93 51.9 55.4 5.S 0.62 Wlrth (1931) 60-80 8.S6-11 5-7 50-100 25-50 37-63 2-10 3-8 O-:©. 5 Av. 30 50 5 6 0.1 Miller (1933) 56F 87,1 IB 6.325 86.16 Thormahlen (1935) 6.0 82.2 30-40 50 7 10 1.0 Zemljic (19S5) 50 6.1169 70.08 S2.4g 52,52 5.79 8.96 0.8 Thijn (1936) 12 46-109.6 3ST54 5QT4B 1-5 6-23 0-1 Bell & Irwin (1938) 6.1828 95. 74 34.89 42.27 9.46 12,4 1.003 •Wlrth & Mader (1938) 5-7 50-100 39 50 5 6 O.X Delaune (1939) 5 6.39 10S.25 25.9 58.1 8.0 7.0 Delaune & Msyhevr (1941) 4 6.05- 95.so 19.1- 56- 7.2- 5- 6.80 lo 7.63 £9.8 61.6 9.0 15 Ferguson ei al (1945) £5F 6.3291 89.1152 34.73 41.S4 7.94 14.87 0.S2 "literature review ••y-female, M-male ***The percent of hemoglobin was conrerted to grams from conTersion tables of Bauscii and Lomb Optie&l Company. 1930 Table 2 Hemograms reported on hoi'ses Hemoglobin Differential rioiint.. ner cent content*** E.B.C. W.B.G. Neutro Lympho Mono Eosino Baso No. of Gtt. per millions hundreds phils cytes cytes phils phils Aiatlior iinimsls % 100 cc. per cu.ima. per cu.mm. % % Burnett* (1917) 68-11 9-11 5.5-10 65.-110 50-70 35-45 1.5-3.5 1.5-4 .2-.7 Eabersang* (19Si) 6.5-9.5 69.-110 50-75 15-45 1.5-14 .5-5.8 0-,7 Hauber (1924) 25^ 21p** 7.8-11.0 67.-110 50-6 E 32-46 1.0-2,2 1.S-2.5 0.5-1.6 Meyer (1924) 60-70 28 3.S 4 0.3 Eoiiaaawa (19S8) 12 7.201 80,68 54.2 38.1 2.6 4.7 0.4 Dremjatsky 139 65-77 9-11 7.8-8.9 70-93 50-60 26-40 2.4-4-5 3.2-8 1.5 et ai. (1989) Scarboroiigli* 70-9S 10-13 7.8 92.6 56.8 30.4 8.5 3.7 0.46 (1931-3S) mrtli (1931) 7-10 70-100 55-65 16-43 0.3-6 2-4 0.1-0.6 Av. 60 35 3 S 0.5 Neser (1923) 45-60 30-45 2-8 5-9 0-1 Stewart (1940) 36 6-7 66.-118 36-72 13-56 1-8 1-28 0-3 Lamarre (1944) 45-60 30-45 2-8 3-9 0-1 V *Literature review **1"-female J p-pregaant ***The percent of Ixemoglobin was converted to grsms from conTersion. tables of Bausch. and Lonib Optlc)^ Company. 1930. Table 3 Hemograms of the cows -ased in this study Hemoglobin content* R.B.C, W.B.C. Differential count, uer cent Animal Gm. per millions hundreds Weutrc- Ljmiplio- Mono- Eosino- Baso- 100 CO. -per eu«inm» 4SS4S 7,545 14.14 46.7 37.3 10.0 6.0 0.0 42V43 5;850 6.72 23.7 55.3 8.7 11.7 0.6 5643 120 16.51 7.940 7.06 19.3 67,3 5.3 7. 7 0.3 5S04S 93 12.81 6.125 6.36 28.7 54.3 8.3 8.0 0,7 52743 78 10.74 6.790 6.86 19.7 64.7 6.0 9,0 0.6 61643 88 12.12 5.2B0 8o08 27.0 64,0 4.3 4,0 0,7 6184S 85 11.7 6.865 4.26 41.67 44.0 4.3 9,7 0.3 62343 67 9.28 4. 775 6.76 40.0 40,4 13.3 5.0 1.3 6S843 63 8.67 4.530 6.82 31.0 52.4 7.3 9,0 0.3 7543 108 14.87 6.900 8.90 83.35 49,7 11.7 14.3 1.0 7743 85 11.7 6.760 10.32 17,33 63.0 4.7 14.0 1.0 71243 83 11.4 6.615 7.14 E7.0 56.7 0.7 14.3 1.3 72143 75 10.33 5.110 7.66 51.0 33.0 2,0 13.3 0.7 72843M SO 12.39 5.880 7.54 37.7 53.3 5.0 3.7 0,3 lisnge 63- 8.67 4.53— 4.S6- 17.3- 33.0- 0,7- 3.7- 0,0- 120 16.51 7.94 14.14 51.0 67.3 13.S 14.3 1.3 Average 86.3 11.87 6.2832 7.7585 31.01 53.17 6.61 9.33 0.68 Ciompany. 192SO. Table 4 Hemograms of the horses used in the study Hemoglobin content* H. B. C. W. B. C. Differential count. Tier cent Aaimal Gffl. per millions hundreds Neutro - Lympho Mono Eosino Baso Humber 100 CO* per eu. imn. per eu. mm. phils cytes cytes phils phils 6943W 53 7,98 B.OSO 8.4 66.7 26.7 3.3 9.0 0.3 &a43b 68 9.36 4.905 clotted 61.0 30.0 4.0 4.0 1.0 5a43w 88 12.13 11.050 clotted 85.0 10.0 3.0 0.0 2.0 'S'943b 58 7,98 8.440 8.7 66.7 31 0.7 1.3 0.3 6943s M** 54 7.55 6.585 8.04 68.0 25,4 1.3 4,3 1.0 8344 SI 80 11.OS 1S.040 9.9£ 43.4 44.0 £.3 9.0 1.3 92944 date lost before recorded 78.3 14.0 3.7 3.3 0,7 Renge 54-88 4.905- 8.34- 43.4- 10.0- 0.7- 0.0- 0.3- 12.040 9.92 85.0 44.0 4.0 9.0 2.0 Average 67.6 9.33 8.546 8.84 6 7.01 25.87 2.61 4.41 0.94 **M-niale castrate * The percent of hemoglobin wss converted to grams from conversion tables of Bstisch sjid Lomb Optical Company, 1930. 24 Bone Marrow Studies Terminology In 191S Ackernecht reiuerked "biet no unity existed in the morphologic and histogenetic classificfition of marroii? cells. In 1944 Osgood end Seemsn were still pleeding for "an approved •Standard Nomenclature for Hematology'." Witii the exception of Osgood and Asteorth (19S7) end Pitts and Pack- hsffi (1939) most authors haTS adhered to s ggserslly uaifoiK. set of terms in describing t3ie bone marrow cells, Unfortimstely the saiae tenu did not always apply to the same type of cell. Ssnchea (1941), MexLuow and Bloom (194S), Israels (1943), and Piney end Hsmilton-Paterson (1944) preferred the term "heeaocytoblast" to "myelohlest" used by laost all others. Mulligan (1941) chose the term ''stem cell" -wsiichi included the -very early cells in oil series. According to Osgood (1957) the monoblast, myeloblast, Ijmplioblsst, plaaaa- blaat and megploblast are distinct types of cells but Jordan (1929) stated; "to differentiate the various "blast" cells is to ignore individnsi differ ences betffaen cells in 8 given series." Stasney end Higgins (1937) used in addition to myoloblest the vford ^leidcoblest" to desigjiate 8 ceil betiveen the myeloblast and the promyelocyte. Mallarme (19S7) adhered to the term "leuko- blast" instead of either myeloblast or hamocytoblast. lost investigators agreed on a "promyelocyte." Wijitrobe (194S) divided the promyelocytes into *•1" and "B" types. The nomenclature followed by the msjority of authors to describe the developing grenulocyte aeries included a myelocjrte, a metemyelo- cyte and the adult granulocyte for the neutrophil, eosinophil end basophil. Reich (1935), Suarez (1936), Kandel snd LeRoy (1939), Scott (1959) and Ysn Loon and Clark (1945) had a "band** neutrophil, Segerdehl (1935), Theddea and Bakalos (1940) end Mulligan (l94l) s "stab" neutrophil end Lichtenstein and ?.5 lordenson (1959) preferred to use "rod® foim instead of "stab" or "bsnd". Doan (1939) end Rhosds and Miller (1958) used the letters "1", "B" and "C" to represent stages in the deYalopment of grsjiulocytes or mj-elocytes. Yegu- da (1936) used the tsEis "non-granuler*' snd "grenular" rayelocjrte to indicste stages in derelopment. Holmes end Broun (1953) carried the Homan numersls II - ? or YI TRith the neutrophils to indicate the number of lohes, ??he technics! sj^pressions designating the red blood cell series may be diYided i3,to t^;o main groups; the one, "proerythroblasts:,"- erytltroblasts," and "normoblasts" accepted by Jb^inkin (1929), Reich (1935), ME-rkoff (1956), Suarez (1936), Ysguda (1936), Vogel, Irf, raid Rosenthal (1937), Coen (1939), iia-vidGon (1941a sM b) and Sanches (1941); the other "bt^tsopMlic," •'polychro matic'* and "orthochrociatic" erythroblasts or normoblesta as used byMellarms- (1937), Kf.ndel snd LeEoy (1959), Scott (1939), Lichtenstein and Hordenson (1939)., Thaddoa and Bakalos (194G), Maximow end Bloom {194£), lintrobe (1942) End Tan Loon and Clark (1943). Japa (1945) divided them into two groups "early" end "late" erythroblasts depending on the cytological appearence of both nucleus and cytoplasm. Almost &11 authors included the lymphocyte, mono cyte, end plasma cell in routine counts. Many involved the jnegsksryocyte end some enumerated the reticulo-endothelial cell or a reticulrai cell. Only Thad- deas end Bakelos (1940) counted monoblests and ptomonocytes, Sabin and Doan (1927) end Doan (1939) included the clasiaatocyte in the cells counted. The exceptions to the above, Osgood (1937) end Pitts and Paokham (1939), used a set of terms not in general use such as "progranulocytes A snd B", and "rhabdocytes", and "lobocytes" for the unsegmented and segmented poly morphonuclears. In the red blood cells series they used ''karyoblasts", ''pro- karyocytes," "keryocytes" and "mstakarybcybes". 86 With ths idee in mind of having as simple a teraiinology as possible snd yet be useful, the follofdng list of terms to specify the cells obsenred in 1)0116 marxoB was decided uponi "Stan cell" - All tha immature cells which could aot be clgssified into any given series; "Erythroblast" - ill cells in the red blood cell series from the youngest that could be identified with that,series to the normoblast, "NoEHioblest" - Those in the red blood cell series that contained hemoglobin in comparable amounts to the adult red blood cells as indicated by similar staining properties,ajsd ooatainiag auoloi» '^romyslocyte" - Young cells ^-ath non-speciilc azurophilic granules. "IbsiEophilio'* and "neutrophilic myelocytes" - Cells in these respective series included the metainyelocytes, juvenile cells end stab, rod or band forms of other authors. "Eosinophil" end "neutrophil** - The adult cells of these granulocyte series, "Basophil" - There were so few basophils that both developing cells and adults of that series were grouped together, lymphocyte" - Any developing cells in this series were included. •Monocyte" - All promonocytes and distinguishable monoblasts were added here, "Plama cell" - If any developing pleaiia cells were observed, this heading in- ^ eluded them. The list fflskes a total of twelve headings end it is not BO formidable that it should discourage anyone from attaapting to make bone marro-w cell counts. According to Bloom (1945) anyone familiar tfith the ino3:5hDlogy of blood cells should have little difficulty in the recognition of bone marrow cells. 27 Myelograms Tables 5 encl 6 indicate the Individual myelogrsms for all the ex perimental animels in this imrestigation. Tables 7 and 8 contain a summsiy of this study and a comparison Tsith similar investigations carried on elseishere. Bescriptiou of cells It is iae-fitable that all the slides will not bs stained exactly alike princip.'ally as e result of variation in the dilution of stain, sad in timing, frequently there srs darker and lighter stained areas on the saaie slide due to the variation in the mixing of tjie buffer with the dye. fainter stained preaarations tend to show the nucleoli more clearly and the nuclear structures may not be maskad by the staining of superimposed granules. Heavi ly stainsd aaears, though the nuclear stmotures may be mailed, show the greai- ules in the cytoplasra well. The younger the coll the more homogeneous the protoplesn of the nu cleus. Nucleoli elso indicete a young call since they disappear as the cell develops, nucleoli are greyish-blue sphericsl or ovoid bodies up to £ miera in diameter. frequently cell nuclei appear like doughnuts but it is only the ends of u- or rod-shsped nuclei touching or overlapping which give this appearance, in this study the slides were exsmined in the following msnnar. A low power preview ma made to get an idea of distribution, staining and sny- thing unusual, A representative field was chosen for the differential counts, measurements, color comparisons and cytologicel details and was explored under oil iiffiaersioE. 28 Table 5 Percentege distribution of the mBrrow cells from the ribs of 14 cows Cell Types *42245 4274S 564S 52045 52748 61643 61843 62543 68843 .7543 Stffia cell 1.4 2.0 3.4 0.8 0.2 0.0 2.2 3.0 2.2 5.2 Erythroblast 15.6 36.2 29.2 20.5 30.2 11.8 42.4 20.0 35.6 37,0 NoimoblEst 21.4 19.8 18,0 13.6 27.8 9.2 28.6 7.2 £0.6 15.4 Total eiytloroid cells (1) 37.0 56.0 47.2 44.2 58.0 21.0 71.0 27.2 57.2 82,4 Promyelocyte 3.0 1.2 0i6 1.4 0.0 1.8 2.0 6.8 1.5 0,8 Neutrophilic • myelocyte 19,8 24.8 16,2 32,0 21.0 28.8 16.4 29,2 17.2 IS.O Neutrophil 9.4 5.0 4,8 4.6 7.2 12.2 1.4 10.0 4.0 8.4 Eosinophilic myelocyte •4.6 5.6 9.2 7.8 3.0 9.0 2.8 9.8 9.0 10.4 Eosinophil 7,6 0,0 1.0 0,6 1.0 2.4 0.0 4.2 0.2 3.0 Basophils {ell) 0.2 0.0 •0.4 1,0 0.8 0,2 0,4 0*4 0,8 0.0 Total myeloid cells (M) 44.6 36.6 32.2 47.4 33.0 54.4 23.0 60.4 32.8 35.6 . Monocyte 0.0 1.2 5.6 5.2 1.2 7.6i' 1.4 1.6 3.4 2,2 Plasma cell 0,8 0,8 0,4 2.0 1.0 0,2 0,4 1.4 0.8 1.0 Lymphocyte 17.2 3.4 11.2 10.4 6.6 16.8 2.0 6.4 3.6 5.6 Megakaryocytes in 300 sq,, mm. £I 41 23 20 0 8 0 121 7 8 Mitoses 3 7 2 4 5 8 5 0 4 9 My eloi d-erythl'oi d ratio 1.2 .65 .68 1.07 .57 2.59 .32 2.22 .57 .68 *A11 female except 72343 s from the ribs of 14 cows 61648 61845 68343 62843 754S 7745 71243 72148 72843 Baage Mesm 0.0 2.2 3.0 2.S ; ^'2 5.0 2.4 2.8 1.4 0,0- 5,0 2,14 1i 11.8 42.4 20.0 36.6 37.0 38,6 30,8 42,8 31.8 11.8-42,8 30,26 9.2 28.6 7.2 20.6 1 15.4 35.6 22.2 27,0 39.2 7,2-39,2 21.69 21,0 71.0 27.2 57.2 ; 52.4 72,2 53.0 69,8 71,0 £1.0-72.2 •52.66 1.8 2.0 6.8 1.6 ; 0.8 0.6 0.6 0.8 0,0 0.0-^ 6.8 1,51 28,8 16.4 29.2 17.2 J 13.0 10.4 15.8 16.2, 10,6 10,4-32.0 19,39 12,2 1.4 10.0 4.0 : 8.4 2.4 6,2 -1.2 3,4 1.2-12,2 5,73 9.0 2.8 9.8 9.0 ^ 10,4 5.0 8,4 6.2 1,8 1.8-10,4 6.69 £.4 0.0 4.8 0.2 ! 3.0 0.2 3,0 0.2 3,6 0,0- 7,6 1,92 0,2 .. 0.4 0.4 0.8 i 0.0 0.0 0.4 0,0 0.2 0.0- l.O' .34 54.4 23.0 60.4 32.8 i 35.6 19,6 34.4 24.6 19.6 19.6-60.4 35.59 7.6 1.4 1.6 3.4 •r 2,2 l.E 4.4 0.2 1,8 0.0- 7.6 2.64 0.2 0.4 1.4 0,8 1,0 0,6 1.0 0.2 0.4 6.2- 2,0 .79 16.8 2.0 6.4 3.6 5,6 1.4 4,8 2.4 5.8 1,4-17.2 6.68 8 0 121 7 • 8 5 8 83 7 0-121 25.14 2 5 0 4 : 9 9 5 11 3 0- 11 4.9 7 2.59 .3?, 2.22 .57 . : .68 ..27 .65 .35 .28 0.27-2.59 Table 6 Percentage distrib-ution of the marrow cells from the ribs of 7 horses 6943Tr 6843b 6a43w 7943b 6943s 8344 92944 Cell Types I- F I- F *M-c M-c P Range Mean Stem cell 0.4 0.4 1.6 3.4 2.0 2.4 1.0 0^4—3i4 1.6 Erythroblast 19i4 s.o 14.0 32.0 £3.6 31.4 IS. 2 8.0-32.0 20.94 Normoblast 24.2 IS.O 5.0 15.6 15.2 13.6 10.4 5.0-24.2 13. 71 Total e3?ytliroia 45.6 £0.0 19.0 47.6 38.8 45.0 28.6 19.0-47.6 34.66 cells {S) Promyelo cyt e 0.0 0.6 5.0 1.3 1.6 3.2 0.6 0i0-5i0 . 1.83 NeutropMlle 26.8 47,8 56.0 26.6 31.6 37,8 40,4 26.g-56i0 38.06 myelocyte Heutrophil 20.2 16.6 9.0 12.6 15.4 1.8 17.6 1,8-20.2 13.31 Eosinophilic 0.8 3.4 0.4 2.6 3.6 2.6 3.0 0.4-3.6 2.34 myelocyte Eosinophil 0.4 O.g 1.0 1.0 0.2 0.2 1.2 0.2-1.2 0.60 Basophils (all) 0.0 1.0 0.8 0.4 0.8 1.0 0.8 0.0-1.0 0.60 TotaJ. mysloid 47.6 69.6 71.6 45.0 53.2 46.6 63,6 45.0-71.6 56,74 cells (M) Monocyte 2.0 4.4 4.8 1.2 1.6 1.8 1.4 l.S-4.8 2.46 Plasma cell 0.3 O.S 0,8 0.8 0.6 0.8 0.0 0,0-0.8 0.63 Lymphocyte 5.6 5.0 2.2 2.0 3.8 3.4 5.4 S.O—5.6 3.91 Megekaarypcytes 0 0 a 0 3 1 0 0-8 1.71 In 300 sq. mm. Mitoses 2 0 0 2 8 1 6 0-8 2.71 Myeloi a-erythroi d 1.09 3.48 3.76 .94 1.37 1.04 2.22 .94-3.76 ratio *M-c-msie csstrete Table 7 ComDsrison of" cow data with other myelOKrams Middle Range Meen aged Old from from Mercato (1941) cow cows Mean H.-iBrre (1943) Raaae Author teble 5 table 5 Heaao cy toblsst 4.35 4.1 4.23 Myeloblast 1.5-4 Stem cell 0.0-5,0 2.14 Erytliroble st 5.7 3.68 3.69 Proerytli3X)bls St g.3 E.4S 2.36 Pronormoblast 0-1.0 BasopMlie erytlm>t)last 11.5 11.95 11.72 Basophilic Total erythroblasts 17.5 18.05 17. 77 nortnoblest 3-7.5 Erythroblast 11.8-42.8 30,26 Polyclirometopliili c erythroblast 13.0 16.8 17.4 Hemoglobin Orth.och3?omatie erytirroblast 19.5 19.0 19.25 containing Total 37.5 35.8 36.65 noimobls st 2 7-55 Noraioblast 7,2-39.2 21,69 Totel erythroid Total ei'ythroid series 55.0 53.85 54.42 cells 21.0-72.2 52.66 Neutrophilic promyelocyte 1.4 1.5 1.45 Promyelocyte 0.5-3 P romy elo cyt e 0.0-6.8 1.51 Neutrophilic myelocyte 5.6 6.1 5.85 Myelocyte 3-9.6 Neutrophilic metamyelocyts 11.5 Ig.l 11.8 Metgany elo cy t e 3-lS.5 Meutrophxli c Total neutrophil series 18.5 19.7 19.10 Stab cells 7.6-18.,1 myelocyte 10,4-32,0 19,39 Segmented neutrophil 6.3-19 Neutrophil 1.2-12.2 5.73 Eosinophilic proiayelocyte 1.5 1.5 1.5 Eosinophilic Eosinophilic myelocyte 5.0 5.0 5.0 myelocyte 1.8-10.4 6.69 Eosinophilic metemylocyte 5oS 5.S7 5.78 Eosinophil 0.0-7,6 1.9S Total eosinophil series 1S.8 11.77 12.28 All bsBophils 0.0-1.0 0.34 Total myeloid cell B 19.6-60.4 35,59 Monoblsst end monocyte 8,35 9.36 8.85 Monocyte 0-1 Monocyte 0.0-7.6 2.64 Plasmacyts 0.5 0.68 0.59 PlsBms cell 0.2-2.0 0,79 Lymphocyte 1.4-17.2 6,68 Mveloid-erythroid ratio 0.08 0.66 f&A M.E. ratio 0.7 M/E .27-2.591 .676(T. Table 8 Comparison of horse date with other myelograms Hjarre and Hjarre Rsnge from Mean fTOm Berthelsen {l938i 10 horses (1943) Author tebla 6 table 6 Myeloblast 1.5-E.5 1.5-5 Stem cell 0.4-3.4 1.6 Promegaloblsst like cells l-£ Pronormoblast 1.5-3 S.5-5.5 Erythroblasts 8.0-32.0 20,94 Basophilic noimoblsst 4.5-9 4.5-9 Normoblasts 5.0-24.2 13.71 Hb. containing normoblasts 40-60 40-60 Total erythroid cells 19.0-47.6 34.66 P3X>myelo oyte 1.0-2.5 1-8.5 Promyelo cyte 0.0-5.0 1.83 I^elocyte 2-5 2-6.7 Metemyelo cyt e 5-15 5-15.7 If ©utrophili c iStab cells 5.5-11 5-10,7 myelocyte 26.2-56 38.06 Segmented neutrophils 6.5-14 6.S-16 Neutrophil 1.8-20.2 13.31 Eosinophilic Bo sinophil 0,5-1.5 myelocyte 0,4-3.6 2.34 Eosinophil 0.2-1.2 0.60 Bs sophll 0—0» 5 All basophils 0,0-1.0 0,60 Total myeloid cells 45.0-71,6 56. 74 Monocyte 0-0.5 0-0,5 Monocytes l.S-4.8 £.46 Plasma cell 0.1-1 Piesma cell 0,0-0,8 .63 Lymphocyte 2-6 Lymphocyte 2.0-5.6 3.91 Myeloid—enrthroid ratio 0„5 M/E -94-3.76 , lf64 32 The two main sources of reference used in iden1:ifying and describing the cells were "Atlas of HsaBtology" by Osgood and Ashworth (1937), end the •Titmsell Book of Cqlor" (1929), "Colour Terminology in Biology" by Dade (1943) was occasionally referred to. lor the purpose of specificity color comparisons were made ivith the Muasell Color book and the approximate color or range of colors was deterained. Comparing transmitted light with reflected ligHt preseated some incongruitieB. The most predominating nuclesr color was chosen for comparison. The nuclesr color of all cells may be epproxtmetely matched in the 5,0 rod-purple color chart. The darkest nucleus Tcith a yelue of 2 and a chroma of 4 (2/4) ms found in the normoblast. The pale staining -which was frec^uently found in the moBio- cyte nucleus may be designated 7/4, Other cell nuclei matched colors lying be tween these two extremes depending on the stage of deTelopraent end the intensity of stsining. Like that of the nucleus the cytoplasmic color varied idth the degree of staining but in addition there was considerably more variation between cells. Frequently the cytoplasm was so pels that it might be designated "colorless". The pale blue of the monocjrte and a color in the erythroblast series compared very favorably with a purple-blue-purple shade designated 10,0 B 8/2, At the other end of the color renge •was the purple^fclue of the stem cells end the young erythroblest, 5,0 purple-blue 4/10 or 5/10, The cytoplasm of the eryth roblast that had begun to take up hemoglobin matched 8/2 in the 5.0 green- yellow Munsell color plete. The nomoblsst cytoplasm compared favorably with 8/2 in the 5,0 yellow-red plete. The color of the extra-nuclesr protoplasm of other cells ranged from 8/S to 5/8 inclusive on the 5,0 purple-blue plete. The various shades of the eosinophil granules may be designated by 5.0 red 8/4, 10,0 red-purple-red 8/6, 5.0 red-purple 7/E, 7/4, 6/2-6/8, 5/6 33 and 5/8. The basophilic granules wldch were intermingled Mth the eosino philic granules were much the same color as tliose of the basophil itsslf. The basophil granules matched colors under the classification 5.0 red-purple 2/4, 2/6, and 3/6 in the Munsell color book. The azurophil granules of the Ijmiphooyte, monocyte and promyslocyt© were about the colors 4/10, and 5/10 in the 5.0 red-purple plete. The iffliallness of the neuti-ophil granules made color comparisons dif ficult but as nearly as could be ascertained they are the color of 8/4 in the lOS red-yellow-red set of hues. The Eodacolor prints in Plates IT - IS only partially represent the abOTe colors. In the psrta where the background is too blue the reds are not typical. The colors illustrated in Plate X compare very favorably ^fith those in the "Munsell Book of Color. Generally speaking there is so little difference between horse and cow bone marrow that the cells need not be described separately. Stem Cell; The steia cells measured varied in size from 12 x 14 to £5 2 30 fflicssi. These included all the young cells vMch could not be classi fied in any definitive series. The reddish-purple nucleus hsd a homogeneous finely reticulated kaiyoplasai with 2 or more pele blue nucleoli. Bloom and Meyer (1944) did not find a nucleolar membrane in the stem cell of dog marrow but the nucleoli in these cells in the horse end cow appeared to have a very definite nucleolar manbrsne in most instencas. The nucleus -was large in rela tion to its cytoplasm, lor exeiaple, one cell measuring 28 z 22 micra had a nucleus 14.5 x 18 micra. Other cells had a narrower rim of cytoplaaa such as a cell measuring 13^5 x 17 micra with a nucleus 10 x 15 micra. The cytoplasm, which had a varying color range from a pale blue to a greyish-sky-blue, often 34 prsseated s mottled appearance end frequently coatsiasd a vacuole or two. Some of tile atem cells contained a fe?r azure staiEing granules in the cyto- pl&Eiti, For illustrations of this ooll refer to Piste lY, ?igs. 1 emd 2 and Plate X. Ervthroblast and Noiaolilast; Davidsori, Davis and Imes (194S} have aptly defined the srythroblast as "any nucleated cell capable of dii'fer- eatiation towards sn erythrocyte" and divided such cells into four groups ia- cludiag the stera cells of that series and the orthrochromatic siythroblast or nqxjtioblast. The erythroblasts, here iaclude only tifo groups, the basophilic erythroblasts and those beginaiag to teke up hemoglobin (the polychromatic erythroblasts of some authors). Tiiose measured varied in siae from 7 x 7 to 15 X SO.micra. One in the process of mitosis •y,'3s 15 x 23 luicre. fhe nucleus varied from the homogeneoue reddish-purple, nucleoli-coateining nucleus of the most primitive erjrthrobl&st to the chromatia-clumped dark purple nucleus of the late erythroblast just prior to complete heaoglobinization of the cy toplasm, The structureless cytopl&sm varied in color frora the blue of the more primitive cells to the steel grey or even greenish~grey of the stage near the normoblast. The noraioblast presented size variations of 5 x 7 or 6 X 6 to 9 X 9 micra. One in mitosis was 12 z 12 mi era. Aacorciing to Marfcli (193L) the erythrocyte of the ox varies in size from 4.4~7,7 micrs Tsith en average of 5.1 and the horse red blood cell varies from 4.0 to 7.5 \ilth en average of 5.6 micre, Conseouently little if .any si20 differences betiveen the developing red cells in the tv/o .species isould be . expected. None were noted; The py?£notic nucleus isas so purple, as to appear eljaost black and was so dense no internal structure could be made out. Israels (1941 a and b) has described the maturation of the erythroblast. The cell shrinks to one half 35 its original size; the cytoplasmic color progresses from basophilic, to poly- chroimtophilic to eosinophilic with the increase in hemoglobin content, and the nucleus shrinks, condenses and finally becomes a dark featureless mass. The nuclear structure changes and hemoglobinization are not necessarily syn chronized into any set pattern for occasionally a small cell with a pyknotic nucleus inay retain quite basophilic cytoplasm while the opposite, a large cell Tdth an erythroblastic nucleus and coinplete hemoglobinization of the cytoplasm, mj occur. Israels (1941a) claimad the latter was associated with some in creased demand for red blood cells and should not be considered normal. For example in horse number 6843b, cells with hemoglobinized cytoplasm were as large as 13 x 14 micra» This animal had the lovrest rod blood cell count (4,905,000) of any of the animals studied. This variation in development need not be a matter for concorn, however, as the total for the red cell series is a sufficient index to the state of the bone marrow. Sndicott and Ott (1945) made a simple classification of marrow-cells in the rat and one grouping was "red cell series" which included all the nucleated precursors of the red blood cells. Illustrations of this series are shown in Plates 17, V, VI, VIII, IX, Pigs. 1 and 2, and Plate X. Promyelocyte; The promyelooj'tes oaried in size from 15 x 16 to 21 X 22 micra. This cell had a spherical or ovoid reddish-purple nucleus with usually 2-5 pale blue nucleoli. The light blue cytoplasm contained small fair ly evenly distributed azurophilic granules. SometijiBs a rim of deeper blue cjrboplasm was present. For illustration see Plates VI, VIII and IX, Fig. 1, and Plate X. leutrophilio myelocyte; The cells grouped in this series included several developmental stages with morphological variations but all had about S6 the soffls staining properties. The size ranged from 10 z 10 to 16 x 18 Jiiicra. The reddish-purple nucleus varied in fona from the spherical "finely chroma- tiaed" auoleus of tho youngest cell in the seriss, through successive stages of oval, kidney-baen-aad u-ahapes to the darker staining, "chromatin-clufflped" nucleus beginning- to show coastrictioxis but not aegmentsd into lobes. These nuclear changeij correspond 'co the myelocyte, metsffiyalocytQ or juvenila, and stai'f cells of other authors. The cytoplaaa varied from colorless to pale blue vdth fine neutrophilic granules. The early stages often reteinad a few larger asurophilic granules. Examples of tiiis cell are ahom in .Plates lY, V and Till ligs. 1 aiid 2, Plate Jl I'ig, i sad Piste I. Eosinophilic ayelocytQ; These cells pess.through the sane davelop- Biaatel stages as the neutrophil. Thsy are easily identified by the eosino philic grenulss which in~the cow are 1 micron or smaller but in the horse vary from 1 microa to 3 Mcra. Because of the auall size of the granules in the cow it wes impossible to count them tut in the horse eosinophil, from 30 to as many as 125 were seen. Hirschfeld (1897) counted from 20 to 40 gran\iles, Hs stated that the largest ones might reach the sise of the goat erythrocyte. The cells measured were as large ss 26 x S6 and 2S x 20 micra in the cow and one in the horse was 23 x 53 micra. In the young cells there was considerable variation in color of the granules, some azurophilic, others taking on a blue-gray shade. Most of the were spherical ..but a few oval ones vjere observed in the horse eosinophil. Hirschfeld (1397) observed these elliptical grenulss too. According to I^oraiey (1915) the • eosinophilic granules are endogenous, being differentiated from ths baso philic protopl&sBi. He i'ound that they changed from atiall basophilic granules to evon larger eosinophilic ones than were to be found in the mature 00sino- 37 pMl. The cytoplasm appeared grey-blue when not obscured by the grsnules. The reddish-purple nucleus changed from spherical to ovoid to u-shaped, be coming lobed la the adult cell. One or more nucleoli were observed in the most immature cells of this series. Turn to Plates I? Figs. 1 end 2, Plate Y Pig, 1, and Plate X for the illustrstiocs; Basophils; The largest immeture basophil messured was 18 x 21 miere and the eaiallest adult call in this series was 11 x 11 micra. The nucleus when discemable ITOS a redfiish-pui^le color but it was frequently masked by the basophilic granules which were a deeper shade then the nucleus. The spherical grenules Taried in size, the largest being about a micron in diam eter. The cytoplaam TOS a pale blue, According to Hirschfeld (1897) the mest cell (basophil) of the horse TOS large end displayed en abundance of granules such as did not occur in any other snimsl. He thought the granules were needle shaped ("nadelfoimig"). No epprecisble difference ooiild be de tected between the size of the basophils of the horse and cow and needle shaped granules were not observed. Staining variations of the granules ranged from a muddy reddish color to a very dark purple with a brownish'csst, Plate Z contains a drawing of a basophilic myelocyte from the cow and Plate IX rigs. 1 end 2 are photomicrographs of horse bone marrow including basophils. Plssma cell; The plesma cell of both the horse and cow was easily distinguished from other cells but its differential characteristics are dif ficult to describe. The blue of its cytoplaaa was just s bit brighter then that of the erythroblast. This and its usually eccentrically pieced nucleus with its perinuclear clear area all combined to set it apart from similarly stained cells in the lymphocytic and red blood cell series. The largest plas ma cell measured was 14 x 19 micra with an 8 x 8 nucleus. A mailer cell 38 (11 X U) had 0 nucleus 7 z 8 mi era. The chroms-fcin netiraric was heavy and eosrse textured in the older cells hut of a finer structure and less dense in more immature cells, Miehels (1931b) has witten e review on the plasma cell including moi^hogeaesls. Plate IV fig. 2 shows an sdult pleema cell in cow. hone marroWi Plate Till Hg, 2 shows one dividing emitotically in horse merrow end Plate X. includes a drawing of e plssmehlast from the horse, Megakeryoovte; Aekemeekt (1912) described two t3rpes of megakaiyo- cjrfces in the horse, one a multinuclested cell, the other a single nucleated cell having somewhat different staining properties. He found trsnsitionel stages between the two. He suggested thet the multinucleated oells might be a regressive product of metamorphosis, Kingsley (1935) made a study of the development of the megiekaryocyte in pig embryos, According to him the mega- karyooytoblast is a small cell (8 miera) end ean be distinguished from the hemocytoblast by specific cytopleamic granules. The cell increases in size, the spherical nucleus becomes oval, then horse-shoe shaped and finally bell shaped. The disappearance of the nucleoli and "vesiculetion" of the nucleus accompanies these changes. Subsequent fievelopmeiit includes increese in size and amount of the cytoplasm and nucleus. It may have been these different stages in development that Ackerneckt observed. Limarzi and Schleicher (1940) included the steps in the maturation of the megakaryocyte in a study of thrombopenic purpuj^. They grouped thea into young, adult and degenerated types, levy (1945) concluded that they are abnoimel oells, an outcome of mslfonned cell divisions and no special function should be given to than, ?/hatever their role, that of platelet formation or merely fxmctionless de velopmental abnormalities, a description follows. The megakaryocyte is the largest cell in marrow. Its size varied from 15 x 21 to 85 x 100 micra. 39 The cells were spherical or cvoid. Some of them had large pseudopodia. (Plate 21 ilgs, 3 aad 4) Tery few were encoiiatered in the horse end msny of those did not seon to have any cytoplasm. la the cow many more vfere ap parent, These presented tivo distinct cytological pictures. One hed s dark "blue to plum colored nucleus mth an area of reddish-purple granular cyto plasm. It the outer edge of the cytoplasm the granules were absent or sparse sad the blue cytoplasai wss apparent, (Plate ZI i'lg. 8) One cell presented the exact opposite picture, a perinuclear area of blue and an outer rim of fine reddish-purple granules. The other type had a less dense, lobed or folded nucleus surroxmded by a cytoplasm filled to the edge %lth tiny reddish- purple grcnules. In some cells the cytoplasm eppeered to frsy out into par ticles resembling platelets. Piste H Fig, 3 best illustrates this. Accord ing to Limarzi and Schleicher (1940) platelets are formed by either a detach ment of portions of the cyiJoplaam or by cytolysis. Schenker (1939) f&Tored platelet fomation thi'ough disintegration of mature megaJcaryocyte plsma and found evidence to support the theory that the platelets are of nuclear origin. Plates Y fig. 1 and XI Fig, 1 show the usual megalceiyocyte. Mitosis Tables 5 and 6 indicate the number of mitotic figures encountered in the process of counting 500 cells. They were chiefly in the red blood cell series. Subsequent examination of the elides revealed cell division in prsc- ticelly all of the types of cells. According to Osgood (19S9) all the "blest" and "pro" cells divide by mitotic division and the more mature erythroblasts, plasmacytes and lymphocytes divide mitotically, Kienle (1943) found that the hemocytoblasts seldom shov/ed mitosis either normally or in leukemia, that the myeloblast exhibited more prophases than any other phase while the promyelo- 40 cytes oeourj-sd freQueBtlj* Is asetepbsss sad tslop'sase. Jeps (i94£'} coasted tk«3 swabsr of ai?iciiag eeli$ per 1000 auolssted cells snd tha pro portions of each stfegs p»r 100 e@lls» He coasidersd tl)(^ .jotikbI awsber of ^iTidlsg cells t-o ba IS pgr 1000 auclasta^ cells, dis?xiW^8^S &s follovrs; 40p propssse, 45$ setsphass, lOf aaftpkess ©»d talopJu^s©; 9^ ssyeloeytea v')a4 ^ ayelobisgtf? coagrlsett 48 altoses .^er 10!) snfi 91^ late god ^ esply eiythrobl&sts sftd« m tM otbap 51^, Piste X fig, ? lllastpftes es exythro- blest iB fflitotie di-?i!iiioa ijrcd Pliste ¥111 Fig, 8 3boi*s s plfisisfi: csll dividing by S5^dt03is. My^ioiS-srrthreld r&tio The Kyeloid-siythrold rstio, She of (^fsveloplag red feloofi cells to tfeoBs iri tlis TKhitfe blood eeli series, is sa ISEgort-erit ladss of tbs activ ity ol' the ssrrow, Fliould & nystmis 4ms,M for ftii Ineresse is ono or tbs other of these ssriss- be roflscted la the msrtovr tfe-e ysusl ?j:itlo s-otild be upset, fbis retio 'Ss?;® 3et«'n-ala®i by «;a 8BS aoxaobleste enS coapfejrtag t}j« flgap® with a st»iiey «aa of t-fe« fevslop- isg grsauloeytst?. These ey# i&SiGst&d la fsfeles 7 enS 8 by "fotfii ssyeloicj cells'' er^S "fotal erythreid cells", fha syelolS-ervtaj'oig retlo, S/S, given iii the 9ff®© tsfeles Ttnlae fim ,£7 to £.2 for tfee eo® sac .94 to S,*!^ ia tfe® tors®, KrecJjs (1941) ststad tli^t tbs aoi^al syelold-srsftbroid r>?tio for aea Tfiries from 1;£ to 156 la Inssltby st^sills. SMie tksre i« considarf-bls v&ri- etioa 6®oBg' laditriduisls tho asaa l/I rsstlo for tfee oows \i8e6 isi thie st'udy V0B g5,53/5g,6& or .676 {saa ^abl®!]), fb© sesa for t'm hoTS&s ms SS,?-!/ 34.6S or i.64. (see TeMe 12). 41 Eoue-hsi'ling process Plalies Sil to I? sfeo's^ tfee Uealiag, proeees thf)t follo'ss tlife -use at the drill in prosuria^ ssrrow from rib of lfe?i fcorse, ffee .eargic«l pro- esfiiir® of obtsirdni? sewow ®as ORxtim out fft tb& iatsnrfels bsfore the f-aimal «ss klUed; 9 ? fisys, B m&ks 2 dsys, S m&ks- £ deys, 4 weeks g fieys, S w»^» 2 d«ye» -2 ise^ks 1 dey, 9 csj3, 4 fipys, g dsys, 6| hours 2 hoars. A% fjiitopsy ths p^rtioa of the rifeg ificlyslng tfe© site of tise SrlU hols W6S obt£in.®d. These wtrs s«ltse%ueBtly fiec»lclfied» «sbed4« |)a.ref.fia rad histologlc&l ssolioss preperefi. ?l&tgE 211 HII aho® coa- sseatiTe gtegss is iJi® ®«riy be&Iiag |700«8S» High po'sar aeplflcetloa sSsoKed ss iBctsfisisig sua&®r of fibrolsi&stg ••stiii. th& leagthsaefi ti?!s iatsrTC-l batesea tl^s operatioa esti kiiliag of the asi^sl, Piste 17 fig. 1 3ho?;S the (irllisd ¥ltb. t-; loose i'lforous eoaasctlOB tlssw®, Flfctes H? lig, g fjR^ II' ?lg. 1 sto"? boaa filling tJis drill site ssad ia thg latter 52S?ii-r»g is prfecticfilly coiaplste. The borty Issseileis eppe&r to ftxt-end pfcrsllel to tije fiiyflctios Gf t&e, drilled hole, JV Fig, 2 sso^fs e loagil!:?jdiaal se«tio» of ft rib which hefi aot beeu dtlile^ iato. By rsvle^ing tiiaeo slisaa it «§- pesrs t&r.t. tha isesllBg procsss pj-oceedis rdoBtj tfes sme tias psttarii fis m ordiaery frsotnrsi. Osily iatrKm®s,bn;.aotis boas fossstlor. obssrvsd, Sfiffi|)iing exi30ri!Q0st A ssjspiiag expariaeEt eoasistsfi of getting Eerros- from t'so poeitio&ss efeout m isefe ftpsri oa eeefe ot tir& ribs- In coi?, fh& ri.h$ the Sth to ISth iiM tfes boSy leTSl ms that indlcstsd in Plftell Hg. I. tffcase d&ts a» si50\i® iri fsble 9. the ae&ft.® sbewi ia fabi® 10 vefm Xtmlrd 8teiilstie£''lly by fta.&lysfls of Terisaee. ?&© rssiits smmd three slgaiflesat dltferaaeoa In th© lEth o US) o ® «0 H O •«S'SD CO <•-40 O« H• • o « rj t>- o ffi o fi o lO O *0 •H <-( H <0 M « 1? ^ CO "* « O «3 O O ^ o0 tf © «! O ® O O ® G o• oe «« «• r~i S ^ r~4 ^ i£*i sv 14 o S3 O 0' <9 H fr- O O «0 i« O 03 « 4-> Hi H « i-'j H I-S ® -54 6 St. CS ft: O ^ O O O O tti s-s to o •# 6^ «•« • • e « oajifl H®-<£>•!?> OH <-t H O ^ r-i « p-4• H• «»4ii' >> 10 *7 f O o a) o lo «>« o 4'^ «i> O > o Hj • «*« *•••«•• • « « « O iS- O (S o H tv o r/ o o»«! '0 H H w «s H H iO fTr trH (fi W 0 0} Ci O « 'tS Table 10 Summary of means from table 9 8 9 10 11 IS UOTer Lower Totel Stem cell ..9 .g .6 .-9 .3 ,68 .48 .58 Erythroblast 18,0 11.6 16.£ 9.9 S.8* 13.08 10.72 11.90 Hoxraoblast 18.5 gO.5 13.6 11.8 10.9 16.48 13.64 15.06 Total eryth- 36.5 Sg.l 29.6 21.7 14.7 29.56 S4.28 £6,92 roid cells** Promyelocyte g.O 1.1 1.0 1.3 .4* 1.36 ,94 1,15 HeutroDhilift" • £4.6 29.3 23.9 26.2 22.6 27.76 26.90 27^33 myelocyte Neutrophil*** 3.0 5.1 8.0 9.3 12.4 6.40 8.74 7,57 Eosinophilic 10.g 11.6 11.2 15.0 10.2 11.68 11.58 11,63 myelocyte Sosinophil 3.0 B.6 3.0 6.5 4.5 3.24 4.60 3,92 Basophil .7 .0 .2 , .4 .4 .48 .20 .34 Total myeloid 43.5 4:9.7 47.3 68i7 50.5 50.90 52.96 51,93 cells Monocyte 2.3 5.3 3.4 3.3 3.9 3.0 4.3 3,65 Plaaaa cell .5 • .s- .8 .6 .0 .72 .12 .42 Lymphocyte 15.3 12.5 18.1 9.2 30.6* 15.12 19.58 17.35 *Signifleant ^^Significant aegetive trend among the rib averages *'**Signific8nt positive trend eanong the rib averages 44 rib and no significant difference between positions on the rib. The total erythroid cells indicsted a significant negative trend from the 8th to the 12th rib and the neutrophils s significsnt positive trend in the same rib order. Sines there was only one snirasl and many of the cell types occurred in such few nutabers, the esperiment should not be given too much weight until farther work cen be done to substantiate it. In the light of these meager data it would seem better to obtain ssmplea anterior to the ISth rib. In view of Taricak's (1935) findings that the sternal and of the rib chaaged from red to yellow marrow earlier then the vertebral end, piobably it is wise to drill the rib at as high a lesel ss possible end still avoid the beck muscles. DISCUSSION Hemetologistg are not agreed on the value of aspiration end, bone marrow smear methods. Dosn and Zerfas {I9S7) suggested conservatism in draw ing deductions because of the limitations of the technic, the fallacy of drawing conclusions from counting so tm of the millions of cells present, and the debated question, of identificabion and classification. Custer (1932) found the cellular state of marrow to vary in different bones, and in differ ent bones at the ssme level in the same snimal, Dameshek (1S35) contended that for exact cytological study, marrow smears could not be excelled. Ac cording to Young and Osgood (1935) the only limitation of aspirated semples is the loss of structural relstionships, Nordenson (1935) claimed that mar- rD"w from several bones in the seme patient was similar in quality end quantity, lilliems (1935), studying the cellular pattern of human marrow at autopsy, ascertained that the differential count.from different bones in the ssme ease TfflBs esseatielly the seme. Jaffe'(1936) reasoned that the simplicity and lack of technical skill required outweighed the disadvantages of the dilution \?ith 45 blood and failure to give en ia situ picture. Dsmeshek, Henstell end Yalen- tine (1937) thought the chief sdTsntege of puncture biopsy, its sisiplieity, TOs exceeded by its inaeeuracy. Helpep (195?) agreed vdth Custer (193S) that boue merrow is not homogeneous gjid thet satples from one psrt of g bone dif fer from those from another pert. Stesaey ead Biggins {19E7) concluded, to the contrary, that there is sufficient uniformity of h&natopoiesis in dog merrow so thet "the apprsissl of the raerrow of any one region vd.ll reveftl Tjhat the trend of its cellular chfiages is elsewhere in the body." A leter work by the sajiie authors substsntiates this view for hurrtm mariw (Stasney e.nd Higgins, 1939). Stofitmeister and Buchmfom (1939) studied the influence of sternal puncture on the circulating blood snd found it hed no effect on cell composition, Gordon (1941) geirs es' disadTenteges of sny aspiration method, trsuma, the feilure to dislodge immature cells, loss, of topographic relationships and the dilution Mth peripheral blood, Mulligsn (1942) ob- trdned a faYoreble corrole.tion by comparing marrow obteined by sternal punc ture end trephine methods. Eeich end Kolb (1942) found by statisticfd snaa- ysis thet qusntitstive determinetions on aspirated aiarrovrssjaples '.vere in accurate. l^stein end Tompkins (1845) would invalidate differential counts msde from smears on the basis of trauma and inadequate distribution of cells. Steinsr (1945) felt thst a merrow sspirstion method might not be satisf&ctorjr in Hodgkin's disease because of the distribution of the lesions in foci. By comparing rasrrov: smears from the ribs and femurs of dogs, ?an Loon snd Glsrk (1945) ceme to the conclusion th£t such preparations were similar in content, Osgood and Sesiaan (1944) pointed out thst eny marrow prep&rstion whether s section, an imprint, or aspirstsd materiel, vdll hsve blood in it beceuse blood is present in the sinusoids end vasculsr chsimels of both nomal End pathologic merrow. According to Schleicher (1944) any kind of ssmpling from 46 as lerge end complex an orgsn as bone marrow is subject to errors of chance. In spite of all tMs controyersial evidence as to its velue bone marrow examinetion is a useful tool to be snployed when indicated just es other technics are, Iccording to Bloom (1945) no hematologicel study is com plete if the bone marroT# is neglected. In reviewing this study and the data preseeted her« certain points may be noted. The few sten cells may be readily espl&ined by referring to some in- •vestigstioiis of Japa {194S) on the mitotic actiTity of human marrow. He found that out of 100 cells in the myelocytic series only 3 were myeloblssts and in ths erythroblsstio systepi 9 ^re^o esrly erythrohlests, Trautsiann (1940) sel dom sm the stem cell in noimel bone msri-ow. young erythroblasfs (the proerythroblasts of some authors) ere encountered because the multiplication of older cells is sufficient to fill the physiologies! need (Jaffe' 1933). Most of the mitotic figures occurred in the erythroblastic series, Japa (1942) fouind 15 per thousand nucleated cells in human marrov^. llie range in the cow was from 0-11 and in the horse 0-8 per 500 cells counted. In Fig. 1 the neutrophilic myelocytes end neutrophils in the marrow were plotted on the same graph ^Ith the blood neutrophils for each cow. The resultsnt curve seems to bear out the statement of Doen and Zerfas (1927) that there is a striking reciprocity betiftfeen the neutrophilic myelocyte and the mature neutrophil. lig, Z presents s similar correlation for the horse but such deductions ere not so eppareiit perhaps because of fewer eniiasls. Curves compering the eosinophilic series ivith the blood eosinophil for both the cow and horse ere shova;.:ia Slgs. 3 and 4. There appears to be 4V a positive correlstion between the eosinophilic niyeloeyte snd the marrow eosinophil but BO interdependence between these end the blood eosinophil. Totteimaa (1956) made sueh a study oa 66 porcons vath •broao, tapevform ftnd concluded that there ms no parallelism betvieen eosinophilia in the blood and in the bone marrow. Barta (1923) found that eosinophils might be in creased in numbers in the bone marrov^ without appearing in the peripheral blood. In similar curves for the red blood cell series (i'lgs. 5 end 6) there is a much mere favorable ccrrelatioa indicating that there is a rela tion bet-weea the erythroblasts, norBiobl&sts and peripheral red blood cells. Individual meturation curves of the neutrophils illustrated in fig. 7 indicate that all fourteen cows shov/ed a siiallsr eu3?ve, A mean of these curves is plotted in Ilg, 8 and compered xvith the data given, by HJerre (1943) end Marcato (1941). Cotti (1939) illustrated s gradually ascending maturation curve for human merrov?. Figure 0 shows tho individual msturetion curves of the aeuti-ophils of the seven horses \ised in this study. They com pare favorably with each other and sre like those of the cattle. TablevS 11 snd 12 compsi-e the peripheral blood with the bone marrow for both the cow and horse. Table 13 suimiiBrizes the bone marrow findings in this study. In the lavSt decade great strides have been made in human medicine in correlating the bone marrow findings and certain diseases. Some of those in vestigations are mentioned here.that they may point the my to similar re searches in the field of veterinary hematology. Dreyfus (1936) valued bone marrow examination in cases of myeloma, lymphoma, acute leukemia, anemias and in atypical Hbdgkins disease. Iccording to van de Merwe (1936) the absence 48 Blood naui "(2713 5Zm5 AN/MM NmBER i'ig. 1. Comperlson of neutrophils of bone marrow and blood of the cow. 49 75 B/ood new/'- 55 50 25 25 /Marrow neut. M3b mjw i/J^3b C./^35 AN/MAL NUMBER Fig. 2, Comparison of neutropMls of bone marrow aad blood of the horse. 50 Blood eos. Eos. mifel. \ jMarroiA/ eos- '""VZ13 ^""7Zm3 miAM NUfAd^R Pig. 3. Comparison of eosinopMls of bone marrow and blood of the cow. El fzf^ AN/FAAL NMBS:/? Hg. 4. Compsrison of eosinophils of bone marrow end blood of horse. 5£ /5 /3 IZ // 10 V 7 L S 4 5 Z / AN/MAI. NutA6CR Jig. 5. Comparison of red blood cell series in the marrovf with the peripheral red blood cell counts in the cowa. ML.RB.C. 4^ 13 /^pfa! ercjlhroidser/es 14 /Z / / \ HZ n % /O M B/ood r.b.c. 32 f 30 2? 8 26 z4 1 frqfhrobhsf Zl Zo (= li 5 H /^or'/Tob/asf' iz /o g •i L W5\N 68^5 b km\N T^45b i.ms S344 9Z?44 ANIMAL NU/ABER I'ig. 6. Coinparison of red blood cell series in the merrow with the peripheral red blood cell counts in. the horses. 54 5o\ HS\ P\ 55\ \SZ043 50 25 \ Z0\ l5\ M , mcrro^ 10. .,x» ueutro^mlB oJ Sig' '• 55 MEM 25 20 promcjel. /^euttvLfel. /Aartvivneut Bhodneuf-. Fig. 8, Maturation curve of the neutrophilsof the cow com pared with that of other investigators. 56 Promuehcufes Neutrophil/ Neuirophils Neutrophils ' mifelocqm marrow blooa lig. 9. Maturation curve for the neutrophils of individual horses. 57 Table 11 Comparison of peripteral blood vdth bone marrow (cow) Periubersl Blood fron.Table 5_. s Bone Marrow « Total $ myeloid • from Table 5 Mimsl leucocytes cells* errthroid i myeloid ervthroid 42243 14,140 52.7 7,545,000 44.6 37.0 42743 6,720 36.0 6,850,000 36.6 56.0 5643 7,060 S7.3 7,940,000 32.2 47,2 52043 6,360 37.2 6,125,000 47.4 44.2 5S743 , 6,860 19.2 6,790,000 33.0 58.0 61643 8,080 31.6 5,280,000 54.4 21.0 61843 4,260 51.6 6,865,000 23.0 71.0 6B34S 6,760 46,3 4,775,000 60.4 27.2 68843 6,820 40.3 4,530,000 38.8 57,2 7543 8,900 38.6 6,900,000 35.6 52.4 7743 10,320 32.3 6,760,000 19.6 72.2 71243 7,140 32.6 6,615,000 34.4 53,0 72145 7,660 65.0 5,110,000 24.6 69.8 72843 Male 7,540 36.7 5,880,000 19,6 71.0 Mean 7,758.5 39.1 6,283,214.2 35.59 52.66 Myeloid-erythroid ratio 35.39/52.66 - .676 ^Neutrophils plus eosinophils plus basophils 58 Table 12 Comparison of peripheral blood mth bone marrov/ (horse) Peripheral Blood from Table 4 Bone Marrow Total White % myeloid Total red fTOm Table 6 Animal Sex blood cells cells** blood cells myeloid ervthroid 6943w I 8,400 8,020,000 47.6 43.6 6842Eb f clotted 66,0 4,905,000 69.6 20,0 684SW clotted 85,2 11,350,000 71.6 19,0 7943b 5' 8, TOO 58.3 8,440,000 45.0 47,6 6943s M-c* B,340 73.S 6,525,000 53. S 38.8 8344 M-c 9,9S0 53,7 12,040,000 46.6 45.0 9S944 I - 8S.5 - 65.6 28.6 Mean 8,840 92.1 8,546,666 56.74 34.66 Myeloid-erythroid ratio 56.74/34.56 - 1.64 *M-c mele castrate **Neutrophils plus eosinophils plus basophils 59 Table 13 Samery of boas marrow data on the cow and horse Cow Hjrse Cells Rsaige Mean Rsnse Mesn StsB cell 0,0- 5,0 2.14 0.4- 3.4 1.6 Irythroblast 11.3-42,8 30,26 8,0-32,0 20,94 Normoblast 7,8-39,2 . 21.69 5,0-24.2 .13.7L Total erythroid cells (1) 21,0-72,2 52,66 19.0-47,6 34.66 Promyelocyte 0,0- 6,8 1.51 0,0- 5,0 1.83 Neut3?ophilic myelocyte 10,4-32,0 19,39 26,2-56.0 38.06 Neutrophil l.S-12,2 5.7S 1.8-20,2 13.31 SosinopMlic myelocyte 1,8-10,4 6,69 0,4- 3,6 2.34 Eosinophil 0,0- 7,6 1.92 0,:£- 1,2 . 0,60 Basophils (all) 0.0- 1,0 0,34 0,0- 1,0 0,60 Total myeloid cells (M) 19,6—60,4 35.59 45,0-71.6 56.74 Monocyte 0,0- 7,6 2,64 1,2- 4,8 2.46 Pleraa cell 0,2- S,0 0,79 0,0-0,8 0.63 Lympliocyte 1.4rl6,8 6,63 2,0- 5,6 3.91 Megakaryocytes in 300 sq, mm. 0-121 25,14 0-8 1,71 Mitoses per 500 cells 0-11 4,9 0-8 2.71 Myeltfid-erythroid ratio M/E 0.27-2.5 0,676 0«94r3® 76 1.64 60 of megaloblasts in tiie marrov? ruled out carcinoma of the stomach. Dsmeshek, Henstell and Yelentine (1937) found eternal puncture to he indicated in per sistent anemia, leukopenia, thrombocytopenia, splenectomy and for e differ ential diagnosis of eplenoraegsly. Jsgic and Klima (1937) thought that a marrow examination was useful in diseases of the reticulo-endothellel system. Vogel, Erf and Bosenthal (1937) detemined that bone marrow studies were of diagnostic importaace in Gaucher's disease, layeloms, leishmaniasis, mslaria, certain leukemias and csrcinomas as well as of confirm.atory,value in a num ber of other blood dyscrssias. Zanaty (1937) thought some workers exagger ated the diagnostic value of bone marrow counts but admitted their value in the study of the morbid anatomy and physiology of the blood forming organs and the effects of treatment. Hardgrove and Tan Hecke (1939) resorted to bone marrow examination in questionable blood dyscrasias end certain blood disorders. . :^es (1939) observed that aplastic anemia and myelosclerosis necessitated bone marrow study, Schmid (1939) made e study of blood snd bone marrow in human undulant fever. He observed "Bang nodules" in the mar row and thought the platelet fonning function of the megakaryocytes V:SB in terfered Mth. fogel and Bassen (1939) used sternal puncture to rule out leukemia in eases of infectious mononucleosis. Jones (1940) studied the bone marrow in hyperthyroidian and hypothyroidism end found an increase in nucleated cells in the former and a decrease in the latter. Limarzi and Schleicher (1940) wrksd on the reaction of peripheral blood and bone marrow in chronic hemmorhsge and essential thrombopenic pui^ura and concluded the.t diagnoses of purpuric states could be more satisfactorily made from bone marrow studies. Davidson (1941b) confiimed diagnoses of csncer, Hodgkins disease, multiple myelomatosis end lipoidoses by finding specific cellular 61 elaaeats in the boas marrow. Gordon (1941) suiumarized the velue of bone mar- ro\'f studies under four hesdings; to aid in the diagnosis of obscure conditions, to confiim diagnoses, to further tha understanding of disease processes eM as a method of bioassey of certain drugs, falconer and Leonard (1941) used marrow technic as a means of gain ing additional infomatloa and suggested its interpretation in the light of other data. They recorameaded brucelloais and bacterial endocarditis as a field for tother study, Beizer, Hall and Giffin (1942) confimed suspected rayelorae by finding myeloma cells ia the bone marrow. Bamesh^ (1942) con sidered bone marraw biopsy an important tool in the differential diagnosis of refractory anemia and pancytopenia. He also ruled out leukemia in cases of thrombopenic purpura and hemolytic anemia by this method. Kienle (194S) distinguished erythroblastosis and erythroleukemia by bone marrow studies. In addition to those hemopoietic disorders already mentioned liPurkel and Bethel (1943) claimed marrov) aspiration from the sternum to be en almost universal practice in disturbances of the reticulo-endothelial system, cer tain infectious diseases and neoplasms of the marrow csTity. Williams (1943) found the megakaryocyte count in primary pneumonia to be more than 5000 cells per cubic millimeter end suggested a definite re lationship between pneumonia and hyperplasia of megakaryocytes. Bloom (1945) is pioneering in the veterinary field in studjring bone marrow in various diseased conditions in the dog. Some of his findings may be noted here: increased rayeloid-e3jythroid ratio in pyometrs, end aregenera- tive anemia; neutrophilic hyperplasia in marrow in pyometra and streptoth- richosis; decreased myeloid-eorythroid ratio and hyperplasia of the erythroid series ia advanced filariasis, and the occurrence of lymphoma cells in the 62 bojae marrow in mElignent lymphoma. If future bone marroTfi studies should lesd to an earlier diagnosis- of obscure blood dyscrasies in our fem animals it TOuld be of considerable economic TElue. Mitchell (1943) suggested that an early diagnosis of lym- phocytoma in cattle would result in a sa-vlng in feed and care. In concluding,the fact may be emphasiged that this is only e freg- ment of the studies that could end need to be made. Further vTOrk may reveal a more refined technic of obtaining marrow from these ardmals, Schleicher (1944) has completed a study on the volumetric pattern of sternol marroiv in men including the fet, plsstna, rayeloia-erythroid r&tio, end erythrocyiie con tent per cubic centimeter. Such a study -would be valuable in animals, Wirth (1938), investigated the reections of the heraatopoietic system of the various domestic enimals in response to bleeding. He found thst it vsried with the species, Ck)uld this varietion have been due to inherent differences in the bone merroT^. No bone marrow studies were msde. The gross changes asso- cisted with age end variations Tdth sex are yet to be vxjrked out for the various species of animsls. Are lymph nodules present in the marrow of any of our domestic enimels? There is a difference of opinion regarding their presence in msn. Jaffe'^ {19S6) gsve the weight of human msrrow in percent of the body weight, 5,4 to. 5.9, and found five/ninths of s. gran of msrrow per grsm of blood, .He further ststed thst half of the bone marrour of en sdult person is active and concluded thEt the weight of the red mBrrow equeled that of the liver. How would such figures for domestic animals compare? The effect of drugs upon bone msrrow of our domestic animals is yet to be determined. In the futtire bone marrow may become ss important e diagnostic agent in the field of veterinary medicine as it no7f is in the realm of human medicine. 63 mmm This study ivas mdertakea to deteimine the noimsl cytological pic ture of the bone marroi^ for the horse and cow. Samples -were obteiaed from fourteen head of cattle and ssTen horses. The ribs were chossn as the site for securing the marrow ia both species. The samples were obtained by drill ing into the rib, inserting a cannula into the drill hole and aspirsting 1-2 cc. of marrow. Peripheral blood samples were taken at the same time. Blood and msrrow smears were msde and stained vdth Osgood*s (1937) modifica tion of liiright* 8 blood stain. A sampling study vfas made at two different levels on each of fire lib 8 (8th-12th) in one cow and the cell counts recorded. An snelysis of variance of the means sho?ved a significant variation from the mean for the erythroblasts, promyelocytes snd lymphocytes in the 12th rib. A significant positive trend was observed from the 8th to ISth rib in the total erythroid cells, and the neutrophils showed a negative trend in the same direction. Since only one animal -was used, more rork along the same line is needed to confirm these data. A study was msde of the healing process of the drill hole ia horse ribs end photomicrographs made to illustrate the progress of repair. Repair of the bone was almost complete in 7 weeks and all external indications had disappeared long before that. Cell counts were made to establish a "noimal" myelogram for the cow and horse. Three hundred cells xvere counted in the differential leucocyte count on the blood smears and 500 cells ^rere enumerated in the differential count on the marroi7 smears. The mitotic figures encountered per 500 cells veve also recorded. The megakaryocytes were counted in & 300 si^uare milli 64 meter area. The myeloid-erythroid ratio was cieteimined. Cjrtological studies were made of the marrow cells end color comparisoas were made with colors in the Munsell (1929) Book of Color. Colored photomicrogrsphs were made of the hone marTOv; &me&rs to illustrate the various types of cells and colored draw ings of the various cells were incorporated into a plate. The rayelogram for the cow (raige end mean ia percent): start cell: 0*0 - 5.0, 2,14; erythroblBst; 11,8 - 4£,8, 20.26; norftioblast; 7,2 - 39.2, 21,69; total erythroid ceils (E): 21.0 - 72,2, 52,66; promyelocyte: 0,0 - 6,8, 1,51; neuti'ophilic mj'elocyte: 10,4 - S2.0, 19,39; neutrophil; 1,2 - 12.2, 5,73; eosinophilic myelocyte: 1,8 - 10.4, 6,69; eosinophil: 0,0 - 7,6, 1,92; ell basophils: 0,0 - l.O, 0»34; total myeloid cells (M): 19,6 - 50,4, 35.59; mono cyte: 0,Q - 7,6, 2,64; pleama cell: 0,2 - 8,0, 0,79; lymphocyte: 1,4 - 16,8, 6,68; megaksryocytea in 300 sq. mm,: 0 - 121, 25,14; mitoses per 500 cells: 0 - 11, 4.9; myeloid-erythroid ratio (M/E): 0,27 - 2,59, 0,676. The myelogrem for the horse (range end mean in percent): stem cell; 0,4 - 3,4, 1,6; ersrthroblast: 8,0 - 32,0, 20,94; normoblast: 5,0 - 24.2, 13.71; total erjrthroid cells (l); 19,0 - 47,6, 34,66; promyelocyte: 0,0 - 5,0, 1,83; neutrophilic layelocyte: 26,S - 56,0, 38,06; neutrophil: 1.8 - 20,2, 13,31; eosinophilic myelocyte: 0,4 - 3.6, 2.34; eosinophil: 0.2 - 1.2, 0,60; all basophils: 0,0 - 1,0, 0.60; total myeloid cells (M); 45.0 - 71.6, 56,74; monocyte: 1,2 - 4,8, 2.46; plaana cell; 0,0 - 0,8, 0,63; lymphocyte: 2,0 - 5,6, 3,91; megakaiyocyte in 300 sq. mm,: 0-8, l,?l; mitoses per 500 cells: 0-8, 2,71; myeloid-erytliroid ratio: 0,94 - 3.76, 1.64. (A table summary of these data may be found on page 59, Table 13). Graphs indicated a positive correlation between the marrow neutro philic myelocyte and the adult neutrophil in the blood but no correlstion be 65 tween the merrow eosinopMlic myelocyte and the eosinopkll in the circulating bloofi. Similarly, graphs comparing the marrow red blood cell series to the erythrocytes in the peripheral blood suggested some correlation though not as striking as the neutrophil or eosinophil. Individual neutrophil curves for ell the enimals were similar. Figures were not available for the horse but the neutrophil curve of the cot? agreed favorably ?d.th those of other in» vestigators in the field. 66 LI'IEMTUEE Cim Ackerknecht, 1. ^ 1912 Beltrsge zur Kenntniss des Msrkes der RohrenknoolieD. beim Pfeide. Arch, 208:396-414. Alexendrov, A. I. 1930 Die Morphologie des Stemimpunktates von Hunden. Eolia Heeia. 41:428-434, Axinkin, M. 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Path. 15: 154-158. Lichtenstein, 1, and Hordenson, N. G. 1939 .Studies on bone marrov; in Bremsturs children. Folis Heem. 67: 155-184. Lim, R. K. S., Sarkar, B. B. snd Brotm, J, P. H. G. 19BS The effect of thyroid feeding on the bone fflsrrow of rabbits. J. Path. Bact. 25: S23-248. Limarzi, L. R, end Schleicher, E. M. 1940 The reectioa of peripheral blood snd bone marrow in chronic hemorrhage end in essential throinbopenic purpura, J. Med. A. 114: 12-18. MacGregor, R. G. S,, Richards, W. and Loh, G. L. 1940 The differential leucocyte count, J. Path. Bact, 51: 337-368, Magath, T-. B., Berkson, J, end Harn, M, 1936 The error of determination of the erythrocyte count, -Am. J, Clin, Path. 5: 568-579, Mallarme, J. 1937 Le myelogrsmme normal et pethologique. Sang. 11: 804-832, Manaugh, E. C. 1940 Biopsy of sternal marrow and its place in clinical hematology. Meu. Biil. U., S. Yeterans Acte, 17: lll-122c Marcato, Arnsldo 1941e II midollo osseo normals nella specie bovina, (Mielogramms, rapporto leucoeritropoistico, curve di maturazione) Nuove ¥et, 19:173-179. Original not seen; abstracted in Jehresb. "Vet, Med 69: 530. 194S. 1941b Bone-marrow in bovine fascioliasis. Nuova Vet. 20; 219-223. 77 Msrkoff, 193S Uie des KEoefeenmerks ^ntoh fiteriislpunktion. Daat. Arefe. f. iciis, Me3. 179J llS-133. Mewrer, F, D, Aas Joass, T. 0. IS43 The- blood plctyre in e5,uiKs iaflGeuzs. %. J. '?6t. Sss. 4: ga7-«54. M'&xissow, K, 1910 UsterEfuohimgeii Iber Blut uad Biricsgewebe; Sis «bryonels Histo gSTseas des IriOofeeia??-rks dsr SeugaUere. Irols. f, sikr. £.ufet, 7S; 1-113. ead. Eiooffi., F. 194.?. textbook of Ms-tology, t, B, S&undera Cc, Phllfcd&lpMfi. Mfcysr, B. sn& farsj'ts, S. 1924 Zar frege- &&r LpsabkBOtclim la ^.-ssaehlicfeea Eaoeiieaffifirk, ?lrch i^rcfe. £5S; 5?4-68f). Mecijgatic, S, 1925 Oatersusbuafea uber des Geislciit das KaoohGJimRi'kas ces Messchen Stsc&r. 79:88-99. Meakia, ¥. iS4S Studies on tfee effect o? tjis is^ikoes'tcsis orosot-laii factor oa tSifj boue ssrroiif. Asst. fiec, 85:40, Serwe, C» f. ?rs d© 1928 Eoaa Ksm?^ stusi«s ie the. elialc, Polls Hseai. S&: 106-114, m-m. Meyer, S. i3g4 Blfi BIttfeorpboiogi® ©liaigsr Baus •«r»6 lefeopetorittsstisre untor plsyslelosiischea sand SE'tbologisciian Sediagiingeu. ?olin E«iffis, SO: 19g-}!g3. Micbeis, S, A, i9Si« SrytliTOpoleals. A crtttcsl re view of the iiterfiture. folic Bs-as. m !?5-i£S. 1931b Tia-? ^i&sss- M eriticel rsTis?^ of its serpbogeaaais, fane tioB, 8Bd derelopsiaat cep&clty Tiafi»r bo nasi eafi ebaoimsl coadi tions. Areh. F«t5n 11;?'?5-73S, filler, ¥. 2'. 19Sg Blood VGlMe (!eter?ainatl08s io cattls. Coraftll Tat, S;"2;K0-c-3 78 Miller, W, T. 1933 Erytiirocyts, leucocyte, end hemoglobin detemiaations on tlie blood of cattle, mth a note on the blood in Johne's Sisease. Report of tlis H. Y. State Vet. College st Cornell UniY. for the year 1932-1955. p. ?l-80. Milm£;n, M., Listengarten, S. end Kurbsnalie??, D, 1934 Etudes herastologiqueg. Seng. 8: 481-522. Mitchell, G. 1. 1940 Hyperplasia of bone rnerrow and osteohs-aetocbromatosis in a yearling steer. J. M. Vet. Med. A. 96: 741. • Mitchell, K. P. 1943 Lymphocytosis in a cow, T/et. Med. 38; 49S. Mouriquand, G., HSTOI, L. end Edel, V, 1944 Le layelogreKEae per ponction femoral ches le cobeye normsl. C. rend. Soc. de Biol. Peris 138; 850-251. Mulligsa, R. M. 1941 Quantitstive studies on the bone msrroT-/ of the dog. Mat. Hee. 79; 101-108. 1942 A compsrison of needle and trephine in study of bone marrov;, im. J. Clin. Psth. 12; Tech. Supp, 6; 43-44. 1945 Qjaeiititstive studies on the blood end bone lasrro?? of aoi-vbom mongrel puppies. Inst. liec. 91; 161-167. lunssll, i. H, 1929 Munsell book of color. Munsell Color Co., Inc. Bcdtimore, Md. Neser, C. P. 1923 The blood of equines. Union, of fi. Africa Dept. of Agr. 9th sna 10th Reports of Direct. Vet, li'd, and Res. p, 479-557. Nettleship, A. 1942 Bone-rasrrow changes produced by specific antibodies. Am. J. Path. 18; 689-698. Heuiaann, E. 1868 Ueber Bedeutung des Knocheniasrkes die Blutbilduag, Centrabl, f. d, med. Wissensch. 6: 689, Nordetisou, K. G, 1935 Studies on bone msrrow from sternal puncture. Bortzells, Ssselte, Stockholm, ?9 Hye, a. S. 1331 Bos® ratvrto'i: ToiyjAs in rsbbits. Prcc. fioc. Isp. Biol. Ssd. g9: U'Zr. OfclKOU, 1945 Uapublisiefi tifite fro® Merg&ret i. Otilsoa, Micbiesn, Sfc te College, l8st Lgasing, Mleiilgsii, Osgood, 1, S. i§;j7 Mojiosytic ISBkesifs, /refe. Xst. M«d, S§! 9a-3Sl. 1933 M-'srroT?' cultures. ^ syraposim on tba blood, p. glS-F;4l. Balwaity of Wlj^coasla Press, Mcdisoa. iS40 A textbook of Iftfeonstory disgao^-is. Sd ed. The Blskistsn Co., Phileoelcfeie. euc, Asfc'jjorth, C, 1937 /itlea of Haaetology, J, I. St«cey, lac. Seu ireaolsco. and B.roaales, 1. £. iSS? Culture of imrnsm astiow. Set&lls of e eia^le method. J. i®. A, 1Q8; 1W2-1796. s.Bd Saes&fiK, i, S. 1944 The cellQlfir eoispositlofi ot aomsl bona a&rroTi- ns ohti-dse^ by starael pascttsra. Fhjsiol, E$r. 24: 46-69. • Hfcsltloa, H. B, fjBfi frotfiea, i, T. 19S1 iN aaifom s:vs'^«si of hsssRtclogic aathocls for use with oxslatsd ^/esQ-ua blcofi. J. Usb. Ciin. lleS. IS; 476-4S1, . Psippeaheiis, A, 1899 Vsrgleicfoeadg Uatersuctongen Sber dia fileaeatsr^? Susfisaese&tsufig tias rotois Inoobsaserkefe* elnigar Ssygetiere (Mebst Begjerkuagaa ®ur Fr5.g8 des segeassltigaii ¥erhcltiii®.5es der t^rsebiaderxes LsuKOsyteaforssK s«eiasnd&r). ¥ireh, Ircb, 157: 19-7$, P&tn, £. 19S4 Morpbolo^isckfe uad KUKsrisclje Hat^rsuctog wber Icc!ch6Bs?:?iiz«3lisffl bei aoisi&lsa weiasea Lscor^torltijssmeuses. iicts -Soead:, ll: 1-4S. Pia«y, A. iSSg The Kufctoay of t}j« ison® mrrow with spsci&l pefar«nc8 to the 61 s- tributlGQ of tbe red atsrros. &rit. Med. J, g;- 79g-79S. 80 Piaey, A. and Hsmilton-Peliersoii, J. L. 1944 Sternal puncture. Grune end Stratton, Inc. New York. Pitts, H. H, end Psclchsm, 1. A. 1939 Hematology of sternal juarrov; lend Tenous blood of pregnant and of nonpregnant Vioraen. Arch. Int. Med. 54;471-482. Plum, C. M. 1943 Zur Granulocytopoiese bei Ratten. J'olla Haem. 67!ll9-124, Plum, P. 1936 Accuracy of hematological counting methods. Acta Med. Scand. 90:342-364. • Potter, J. S. end Ward, E. 1. 1940 The deYelopmeat of the megakaryocyte In adult mice. Mat. Reo. 77j 77-85. Eeich, G. 1935 A study of the diagnostic value of stemsl puncture in clinical hematology. M. J. Med. Sci. 189:515-520. 1944 Personal conimunication. Carl Reich, 19 East 80th Street, N. Y. - end Kolb, E. M. 1942 A quantitative study of the variations in multiple sternal marro-?? smplQS taken simultsneously. Am, J. Medi Sci. 204; 496-504. . Swirsky, M., and Snith, D. 1944 Sternal bone marrow in the aged, J, Lsh. Clia. Med. 29: 508-509. Ehoades, C. P. sad Miller, B. K, 1938 Histology of the bone mariw in splastic anemia. Arch. Psth, 26: 648-663. Eichter, M. N. 1938 leucatiiE. In DoTmey, Hal, ed. H&ndbook of Hematology. 4:2885- 3035. Paul B. Eoeber, Inc. lew York. Eingoen, A, R. 1921 The origin of the eosinophil Isucocytes of mammals. Folia Haem. 27: 10-68, I?iser, W. H. 1943 Infectious psnloukopenie of cats. K. Am. ?et, 24: 293-299, Eohr. E. and Hefter, E. 1937 Untersuchungen fiber postmortsle Yeranderungen des manschlichea Knochenmaiie. lolia HESH. 58: 38-50. 81 Bg.bi2» F, B. 19SS- OP. the origia of the cells of tke blooa. Psysioi* Rev. gJ S8-69, 192S Boas Biarropi. PhySioi. Rey. 8: I91-E44, ISo&a, G, it, 192? BOKB aerrow es SH oi'GSE. ??oe. Soo. Ex^. Biol, M§4. g&: fijid Millar, i. S. 19'^B iionaal boae a^Trov-. Ie losasy, ifcl, «d, Hccabook of Hesiatolog?. 3: ITSl-i^^aS. Pftiil B. Ho^ber, I«e. Hew Yo2^. . Miller, 5> K., K, C., Tfeoms®, H. M,. g.a4 Kimsl, L. I. i9S& Chf-jsgss ia tfee boJie siarro^'? end "oloofl csllzs of devsloj?iag robblts, J. 'isp, MM. 64; 97-120. SsacM'K, "S. L, 1941 iSfitudio eliHico d« la modtsla oses. HeaieiBS^ Mei., Si:£6§-S0S. Scarborough, n. l§31-2g ihe blood pictyy® of sotsgI Isbor^tory t?niE5/;is, Ifde J. Bioi. Me«S. S: 63-82',' 168-179, 267-B82, 559-573, 4S1-440, 547-55E; 4;69-t5g, 1S9-206, 2gS-344. . SciieJiker, ?. 193S Ober die plIttoIsacMlfieade iusktioa dar MegaJcaryoeytsxt. lolie H&c-ffi. 63; £23-247. Schilling, Viktor 1985 E&s Kaochsfflssa-k EIS Ojrgsa. Beut. SeS. ichnscbr. S1JE61-£64, 244-348, 4S7-469, 516-518, $98-500. ScJilslslS-Qr, M. AHS vol'iuaetrie pattern of sspirstec aorasi hussJia s-tern!'^! ssftrrov.* of sfdes 18 to 40 years. M, Ciic, Peth, 14; STO-S^S. SchRld, M, i93S Blui sad faocijeisjserk bei Morbus B&ag, Scfessiz. MoS.. i^^ckischr, 63; 191-19S. Scott, K, B, 1359 SisrBcl pBJtictyre ia the dl€;:saosis of aiesseee? of the blood fcraiag orgsss, J, Mm. B: 1S7-17K. Segercehl, E. 1955 Sber SternelpualctioaQn. Acts Med, Smncl, 64; I-ISS, 3ergesl, I., Eoaatlsa, 7i„, Parrot, L,, Lsstofysrd, f, snc Chcrpia, A, ISSS la fo»«l® Isucoeyteire ay fteatg dss bovias, e I'etet norssfil et dims quslcixses 'airoplftsaosffie. Arefe, last, Ps-sstror d'Mg^sris 7; 2-xl. 82 Seyferth, C, 1923 Die Stemiaatrepanation, sine einfache Methode zur diagnostiseliea Entnalme von Knochenmerk bei Lebenden. Ceut, Med, Ichnscter. 49: 180-181. Shukers, C, I., Lengston, W. C. and Day, P, L. 1938 Tbe normal blood picture of" the young rhesus monkey. Folis Hem 60J 416-424. SiBson, S. end Grossman, J. D. 1938 The anatomy of the domestic snimElB. W, B. Saunders Company, Philsdelphia, Snith, C. end Hastings, J". 1935 A study of the megskaryocyte end blood pletelet in the rst, Mat. Bee, 64: 95, Stasney, J. and Feldman, W, H. 1958 Leukesoic lyraphoblestoraa in a calf; a hematologic and histologic study. J, Cancer 34: 240-247, • and Higgins, G, M, 1935 A quantitatiTe cytologic study of the bone marrow of the adult albino rat. Anat. Eec. 53: 77-89. 1936 The bone marrow in the monkey (Mscacus rhesus), Anat. Rec, 67; 219-231. 1937 A quentitetiTe cytologic study of the bone marrow of the adult dog, hi. J. Med. Sci, 193: 462-470, ______and ______1939 A cytologic study of the marrow in the fist bones of man. folia Eaem. 61: 334-344. Steiner, P. E. 1S43 Ebdgkin's disease; the incidence, distribution, nature snd possi ble significance of the lymphogranuloraatous lesions in the bone marrow; a review VJith original data. Arch. Path, 36: 627-637. Stewart, J. T. end Holmani, H. H, 1940 The blood picture of the horse, ¥et. Ree. 52: 157-165. Stodtmeister, R. end Buchmann, P. 1939 Beeinflussung der Blutbildung durch die Sternslpunktion, lolia Heem, 61: 312-316, Strieker, S, 1870 Manual of human snd comparative histology, p, 42S. The Neif,' Sydenhsm Society, London, 83 Sundbergj D. and Jkimsy, E. 1948 Comparison of lymphoid cells of bone marrow end lymph, nodes of rabbits and guinea pigs. hi. J. Amti 70; 4S5-499. Suarez, R. M. 1936 Comparative study of stemsl marrow aspirated during life snd .venous blood. Bol. Assoc. med. de Puerto Hico« 28: 87-93. . DiEZ-Rivere, R. S. and Hernandez-Morales, F. 1943 Aspirated bone marrow studies in normal Macaeus rhesus monkeys, iiji. J. Med. Sci. 205: 581-586. Thaddea, S, und Bakalos, D, 194fl Seitrage EUr Stemalpunktion. Folia Heem, 63: ^l--450. TMjn, J. 1. 1936 Over een, naar asnleidung vsn likzucht, ingesteld morphologisch bloedonderzoek bij het gezonde en het zieke rund. Van Gorcum and Comp., N, ¥. Assen, Thomson, M. L. 1944 Changes in the merrow smear in early megaloblestic hyperplasia, Lfincet, London 247: 688-689, Thoimehlen, A, 1935 Beitrage zur Kenntnis dea Blutbildes uns der Biutzusammensetzung bel der Binder Leukose unter Besonderer Beruchsichtigung des STomalen Blutbildes beim Rinde. Ineug. Dies., Berlin... Original not seen; abstracted in Yet. Bui. 6; 610, 1936, Tkachenko, A, ?. 1940 The •morphology of erythroblasts and Tnyeloblasts in norEsl horse bone marrow. Trud. Troitsk. Vet. Inst, Ho, 3, p. 171-198, Tollner, A, 1931 Beitrag zur vergleichenden pathologischen Anatoraie der Lymphade- nose {Lyin.phoz3rtoinatose) und der lymphossrkomatoae des Rindes, Inaug, Diss., Hrsnnover, Totteiman, G* 1936 Contribution to the knowledge on the relation between bone marrow end blood eosinophilia. Acts, Med. Seand. Suppl. 78: 201-806, Trautman, F, 1940 Suprarttslfarbung sn Stemalpunktaten. JTolia Ea®ii. 63: 325-SS7. Turicel, H, and Bethell, I, H. 1943 Biopsy of bone marrow pffrfomed by a new end simple instrument, I. Lab, Clin. Med. 28: 1246-1851. 84 ?8a Loon, Ei J, end Clr-rk, B, B, 194S Eamatology of the peripheral blood, snd bone marrow of the dog. J. Lsb. Clin. Med/gSs 1575-1579, ^ ?aricBlc, T. D. 1935 Zur KenntniB des Markes der Eumpfknocheni. Untersuchimgen zwecks kliniseher Ausvrertung an Pferd, Rind, Schwein, Bind und Ketze, Arch, wiss, u. prakt. Tierheilk. 73:461-475. •Vogel, P., Srf, L, A, and Rosenthal, W, 1937 Hsaatologic ohservatione on bone merrow obtained by sternal puncture, ik. J. Clin, Psth. 7: 436-447, 498-515. and' Bessen, I, A. 1939 Sternal marrow in children in nonasl and pathologic states, M. J. Dls, Child, 57: S45-S68, Williams, R, J. ' 1935 Studies oh the cellular pattern of bone merrow at routine autopsy. Am, J. Psth. 11: 868-870, 1939 The lymphoid nodules of human bone marrow, im. J. Psth. 15: 377-384. 1943 Hyperplasia of megakaryocytes in pneumonia end its relationship to' leukoblsstic hyperplasia of bone marrov?. Am. J, Peth. 18: 1105-11S7. Wilson, T. 1. 1942 The bone marrow in anemia, Med. J. Australia 1: 513-526. 1944 The thyreoid gland and haemopoiesis. Med. J, Australia 1: 261-269, Windle, W. f., Sweet, M. and Whitehead, W, H, 1940 Some aspects of prenatal and postnatsl development of the blood in the cat, inat. Rec, 78; 381-332, Wintrobe, M. M. 1942 Clinical Hematology. Lea & B'ebiger, Philadelphia. Wirth, D. 1931 Grundlagen einer klinischen Hametologie der Haustiere. Urban e^id Schwerzenberg, Berlin. 1938 Die besondere Resktionsweise der hsnatopoetischen Organsysteme bei unseren Hsusslugetierarten. Thirteenth International ¥et. Congress 1: 273-281. 85 Wirth, D, and Mader, M. 1938 Studien zur artapezifischea Reaktlon der Hamatopoetischen Orgensystane (n, Bind). Jolia. Hsem. 61; 9-14. Wiseaan, B. K. 1934 The origin of the white blood cells. J. As. Med. A, lOS; 1524-152:9. Wittraaim, F. 1929 Die klinisohe Bodeutuag der Hmatologie. Berlin tierarztl. Wclmschr. 45: 399-40£o Wolff, A. 1903 Ueber eine Methode zur Untersucbuag des lebender Knochenmsrks von 'ihierea und uter das Beft-egungsvermogea der Myelocyten, Deut, Med, Wchnschr. lOs 155-167, Wolff, E. K. 1933 KQCchbDinark. In Eirschfeld, H. sad HittmGir, 1., eds. Haad- buoh der ellgemeinen. Ham&tologie. Band I, zweite Halfe p. 1089-11130, Urban sad Schwarzenberg, Berlin. Yaguda, A, 19S6 The bone merrov,' in leukemia, J. Med, Soc, Kev? Jersey . 33: 705-?ll. Yoffey, J, M, sad Paraell, J. 1944 The lymphocyte content of rabbit bone marrow, J. Aast,, London 78! 109-112, Young, S, H, and Osgood, E, E, 1935 SterneJ. marrow aspirated during life. Cytology In health aM in disease. Arch, Int. Med. 55: 186-203, Zansty, A, F, 193? Examination of the bone marrovj by gternal puncture. Ltmcet, London 233: 958-962, Zeraljic, I, 1935 0 uormalnt kr-mi sliki gofeda. Vet, Arhiv, 5; 10-29, Zuntz, N., Loewy, A. 1906 Hohenklima und Bergwanderungen in ihrer Wirkung aus den Measchen, Irgebhisse experimenteller Forschangen in Hochgebirge und Lsboratorium. Bong and Co,, Berlin, ACKNOaEKSfflTS 'fhe author is indehted to Er, H, L. Foust for the initiation of this project and his eaeoursgemsnt tbrcughout to its completion; to Dr. G. 3, J'ovder for his iuterest and eifi in. procuring specimens end to Br. I. E. Walsh (deceaaed) for the use of the cettle belonging to the Department of Obstetrics, ^pprecietion is due Dr. R. A. Bunnells of ths Aastomy De partment at Mic'nigan State College for granting the writer time to com plete the problem after joining his staff in 1945; Dr. R* I'. Lengham, Animsl Pathology Department, Michigan State College for sid in taking the photographs; C. S. Brysii of the Department of Surgery end Medicine, Michi gan State College for the uss of horses; Dr. V«. D. Baten of the Mathematics Department at Michigsn State College for the stetistics, and Dr. E. E. Os good and Miss Eve Padtham of the Di'Vision of Experimental Medicine, Uni versity of Oregon Msdicel School for their time in helping mth cell ideati fication and technics. Gratitude is also expressed to the l07ia St&to Colle Library staff and to Dr. Robert Orr in particular; to Dr. R. G. litch of Michigen State College Library and to Miss Sue Biethan, Medical Librerisn at the University of Michigsn Medicel Libraiy. Piste I Plates I and II are photogrsphs of plates in Toltime I end II of ELIeaberger, Ba-um and Dittrich's (1932) "Handbuch der Metoraie der Tiere", lig. 1. Laterel Yiexf of the Jaorse showing the general eres la wMch to drill into the ribs for marrow seicples. fig. 2. Lateral -new, of the horBe with skin snd superficial fascia remoired to sho\f what portion of the Istersl surfsee of the ribs is relatirely exposed, . M.lUj Plate II Fig. 1, Lateral view of the eow showing the general erea in •which to-dxHl into the rihs for marro? samples, lig. 2, Lateral viev; of the cow with ^in end superficial fascia remoTed to show what portion of the lateral surface of the ribs is relatively exposed. 90 Plate II Plate III Materials used ia drilling for bone marrow. 98 Plate III Plate IV Pistes IV, V, VI, VII, VIII, and IX are Kodecolor prints mede from Isstmsn Kodscolor roll film. The pictures were taken vdth a Voightlander csinera supported over s reseerch. Kiicroscope. A lOx ocular and en oil immersion objeetiTB were used. By measurement of the prints the insgnification of the cells WBB deteirained to he approximately TOOz. Since Kodacolor is designed for use in daylight snd it was used here -with artificial light, the color bslence ms not what it should ha-je been. Tbi.s resulted in the general blue- green color of the pictures ead the failure of the reds to shOTj too ?;ell. Plate I more nearly shows the true color. Several prints from each animal axe included to get as vriLde a variety of cells as possible. Cow bone marrow 1. Stem cells 2. Eiythroblasts 3. Sonaoblast shedding its nucleus 4. Neutrophilic myelocytes 5. Neutrophils 6. Eosinophilic myelocytes 7. Monocyte 8. Irythroblast in mitosis 9. Degenerated or sjaear cell. Cow bone marrov/ 1. Stem cells 2. Srythroblasts 3. Nomoblsst shedding its nucleus 4. Neutrophilic myelocytes 5. HeutTOphils 6.. Eosinophilie•myelocytes 7. Monocyte 8. Plasma cell 9^ Plate n Fig. 2 Plate 7 Fig. 1. Go-vf boae marroTr 1. Erythroblasts 2. Hoimoblasts 3. neutrophilic myelocytes 4. Eosinophilic myelocytes 5. Megekeryocyte Jig. S. Cow bone marrow 1. ii^rythroblasts £. Normoblsst 3. Neutrophilic myelocyte 4. Monocyte 5. Degenerated cell 96 Plate V Fig:.. 1 IMlliii Fig. 2 Plate Jl Jilg. 1. Cow bone marrow 1. Promyelocyte 2. UeuttropMlio myelocyte 3. Srytliroblast 4. Komoblast 5. Lympliocyte fig. 2. Horse bone marrow 1, SuocessiYe stages of erythroblssts 2. Noimoblast 2. Neutrophil 98 Plate VI j&X -X -'••^/He, ' '.,' f-' . •< 1 Pig» 2 Plate TEI Cow bone marrow 1, Eosiaophilic myelocytes 2, Nticleus of B noimoblast 3, Seutropbilie myelocyte 4, Plssaia cell 5, DegeaerEting eosinophilic inyelocjrfce 100 Plate VII ^ 'i'\ ' -ii ^ n * Plat© ¥111 Fig, 1, Horse bone marrow 1. Promyelocyte S. Promyelocyte 5. Eosinopldl 4. Srythroblast 5. leutrophilic myeloci'te 6. Lymphocyte (nucleus sixowing bizerrs clover lesf form) 7. Promyelocyte (cytoplasmic granules not in focus) ?ig. 2. Borse bone marrow 1. Plesma cell (amitotic diTision) 2, Lympliooyte 5. Erythroblast 4. Neutrophilic"myelocyte 5. Neutrophils 102 Plate VIII 1 'i • * vV^fn:- "•> f? ' ^-.'T'" '' '• - Fig. 2 Plate IX Sig. 1, Horse bono mrrow 1, Promyelocjrte 2. Elrythroblests 3. Eosinophilic myelocytes 4, Basophil 5. Lympiioeyte 6, Weuifcropliil ilg, 8. Horse bone marrow 1. Basophil 2. Normoblast 3. Eosinophil lOi; Plate,IX Fig. 2 Plate X BraTSings of the bone marrow cells of the horse snd cow. 1. Ste® cell - horse - 15 z: 18 mi era 2. Erjrthrdbiast, cow - 8,5 s: 9.5 3. Late, e'ry-throhlast, cov; -7x8 4. NormohlMt shedfiing its nucleus, cow - 7.5 x 8,5 5. Plesnisblast, horse - 20 z 18.5 6. /Promyelocyte, horse - 16 x 17.5 7. Irythroblast in mitosis, cow - 17 x 19 8. Monolilaat, horse «• 15 x 16.5 9. Neutrophilic myelocyte, horse » 13 2 15 10. Eosinophilic myelocyte, cow - 22.5 I 2S n. Lymphoblast, covf - 10 x 12 12. Bssophili.c myelocyte, horse - 16.5 X 20 IS. Sosinophil, horse - 13 x 14 106 Plate X I .| I Plate 21 I'ig, 1. Megekaryocjrfce (50 x 55 laicre) x 800 Ifote the ragged edge with fragments (platelets) along the border. Hg. 2. Megakaryocytes (50 x 60 mi era) z 760 i. Bels-blue staiaed outer rim of cytoplsBm. 2; Deaser staining purple central mass, •3. Only apparent nuclear mass in the group, ' ,4. Frejgmenting edge. Fig. 3, Megakarjrocyte vdth pseudopods (55 ,x 100 mlcre) x 1200 This cell had a blue steining center with m outer rim of fine azure granules. Fig, 4. Megakaryocyte with pseudopods (5g x 115 micrsj x 800 108 Plate n Plate 211 Plates XEI to XV illustrate tlie heeling process of the drill hole in the rib of the-horse. ilg, 1, Bib drilled 2 hours \efore killing the animal. Ce 30 z. Fibrin in the blood clotj lig, 2. Hib drilled 6|- hours before killing the saimal. Ca 30 X fibroblasts v;ere beginning to appear in the raass of blood sad fibrin. 110 Plate ni vwir<4^ lig. 2 Pl«te mi Hg. 1. Sib drilled £' bsfora the salsE;! killefi, Ce W X. ItJEsbers of filjro&lasts 'rare iJaaresas^, fig. E. Sife drilled 4. fej's feefor® ts,® scissftl •®Rg killsfi. Ce SO ,x. Fi^jrobleats still iaereeslsg m& el^t h&glmlag to fe® rsssrbac. 11£ Plate nil m •lis, 1 iZ^ \ >0m ilg. 2 Plate 117 ilg. 1. Eib drilled 2 weeks and 1 day before killing the enimal. Ca 30 x Provisional callus foimed. Pig, 2.. Rib drilled 5 weeks 2 days before the enimal was killed. Cs 20 x Drill-hole beginning to be filled Td,th new bone. .114 Plete ny VJ«a" Plate VJ Hg. 1. Rib drilled ? ^veeks 2 days before the animal wan killed, Ca SO 2 The periosteal surface of the drill hole precticslly he&led 'O-ver, A longitudiaal section of a rib. which had not been sub jected to the drill, Ca 20 x 116 Plate Hg. g